## 3 Results and Discussion
### The Variation of Micro- and Macro-Elements within AD
#### 3.2.2 Content of Micro-Elements  
Despite the low concentrations of trace metals, they play a vital role in the AD process. Some of them, such as Ni, Fe and Mn, are crucial to the optimal growth of microbes and related methane production. On the other hand, micro-elements such as Zn, Cr, Pb and Cu can disturb enzymatic activities, leading even to process failure [60]. It should be pointed out that sole SS contains certain amounts of HMs. Therefore, co-substrates that indicate low levels of inhibitory compounds as well as dilution or/and absorption properties are of high interest [1]. In this regard, the use of food industrial by-products may be an interesting option. The highest contents of all of the analyzed trace metals in feedstock were found in the control reactor that was supplied only with SS (Table 4). As compared to other SS obtained from the municipal WWTP, the applied material was characterized by high contents of Ni, Cu and Cr, and a relatively low level of Mn [28]. This finding was related to the characteristics of municipal wastewater influenced by the type of industry located in the settlement. The introduction of co-substrates led to feedstock dilution, understood as reduced concentrations of analyzed trace elements in the co-digestion series. This mechanism has been proven by many researchers [1,2]. In almost all of the cases, the lowest metal concentration was observed in the three-component experiments (R6-R7). The major decreases varying between 54-60% were found in relation to Fe. The significant drops exceeding 40% were also observed in the three-component experiments in the case of Pb, Cd and Cu. With regard to Hg, a decrease exceeding 50% occurred in R8, which was supplied with the highest dose of PL and BSG. The reduced content of HMs in the co-substrates was particularly beneficial in terms of AD, which is a highly complicated and sensitive process involving diverse groups of microorganisms [29]. HMs have been recognized as a substantial factor that influences the anaerobic activities of bacteria. Depending on their form and concentration, their effect on AD can be stimulatory, inhibitory, or even toxic [4]. In all of the series R1-R8 within AD, the concentrations of all analyzed trace metals increased as compared to those in the feedstock composition. This fact is associated with bioaccumulation and biosorption mechanisms [61]. The former is an active process that involves intracellular accumulation of metal, whereas in the latter--which is a passive process--metals are bound with the cellular surface. Besides the bioaccumulation and biosorption of heavy metals in the bodies of bacteria, heavy metals may also be adsorbed by an organic absorbent present in the sludge [62, 63, 64, 65]. As shown in Figure 3, this effect was different in the individual series. The highest accumulation of trace metals in digestate was noticed in R6, which was accompanied by decreased biogas/methane production. This fact may be related with the differences between microbial consortia in the reactors. Moreover, the effectiveness of both bioaccumulation and biosorption depends on the specific conditions in reactors, such as the pH or the presence of other competing ions [66]. However, further research should be conducted in this area. As compared to the control (SS mono-digestion), in almost all of the AcoD series, enhanced contents of all analyzed metals in digestate were obtained. There was one exception concerning series R7, for which the concentration of all metals visibly decreased, similarly as in the case of some of its macro-nutrients. This observation might also confirm the synergetic effect that occurred in the presence of highest dose of PL and BSG. Generally, the digestate concentrations of HMs were established at favorable levels for biogas production [67, 68, 69]. This did not apply to Fe and Zn. The former is used by methanogenic bacteria in the conversion of CO\({}_{2}\) to CH\({}_{4}\); it functions both as an electron acceptor and donor [70].