## 3 Results
### Anaerobic Digestion of Microcalga Chlorella Protothecoides
In this study, considering that lysozyme was the only enzyme tested, and the worst results were obtained for that assay (HPEnz), it appears that the effect of the enzymatic action on the substrate was advantageous since the process required less time, about 20 days, to supply an identical biogas volume to that recorded in H units at the end (Figure 1a). As for the volume of collected methane under the different test conditions (Figure 1b), there is a behavior quite like that observed during the biogas production, except for HExt. The fact that HExt had limited methane production at the beginning of the experiment (Figure 1) can be understood as an adjustment of the balance of the microbial population to the absence of lipids into which, only after about 10 days, the archaea were able to convert the intermediate products in the methane. The superiority in biogas volume of HExt over HPA was not confirmed for the resulting methane as, in the experiment's final period, the same volume (115 mL CH\({}_{4}\)) was obtained in both trial units. The volume of gas recorded in the inoculum units (I) resulted from some traces of degradable material mixed with the used sludge, which demonstrated the activity and ability of the microorganisms present in this sludge to be efficiently used as inoculum in the AD tests. The quality of the respective inoculum is further confirmed by the fact that no lag phase was evidenced at the experiment startup (Figure 1a,b) as well as by the positive evolution in the methane amount recorded over time, and, consequently, any limitation observed throughout the different assays' conditions cannot be related to the quality of the applied inoculum. The differences found in terms of gas production between units digesting whole autotrophic and heterotrophic algae (AA and H) are confirmed in the highest removals of the organic loads by AA compared with H (53 vs. 23% COD, 67 vs. 39% VS). Although they started from similar organic matter concentrations in the two assays (32-33 g/L COD, 9-10 g/L VS, Table 2), AA units presented a much more easily digested and converted material than H and were therefore more suitable for the AD process. As for the pretreated heterotrophic algae, the highest COD removal (53%, HPA) was obtained with the autoclave method in the unit that presented the highest biogas/methane production (Table 2). Likewise, it was confirmed that the lowest removal (14%, HPEnz) was obtained for the enzymatic pretreatment, corresponding with the lowest gas production (Figure 1). Consequently, the decrease in gas in the HPEnz experiment can be understood as a disruption process that may be associated with a change in AD bacterial consortium and/or with the ineffectiveness of the enzymatic pretreatment on the algae cells in releasing their content. The pH of the solutions before the anaerobic process (Table 2) is higher than the initial values obtained in the substrates (Table 1). Values closer to neutrality were obtained in all digesters with substrates due to the addition of sludge for inoculation. There were no major changes in the pH values after digestion; however, there was a slight decrease in the values at the end of the experiment, except for HPEnz where a slight rise was detected. Although HPEnz presents a good methane yield per COD removed (440 L CH4/kg CODr, Table 3), the gas production was only observed in the first 25-day period of the test, probably due to a microbial population imbalance that prevented the conversion of the remaining load. This result is confirmed by the lowest COD removal (14%, Table 2) obtained for the enzymatic pretreatment. The comparison of methane yields from whole autotrophic and heterotrophic algae revealed that the former (AA) was more effective in relation to the supplied food than H. However, when the removal of organic matter was considered, the situation was inverted, meaning that H was more efficient than AA in converting the available material to methane. Examining the H behavior in relation to the gas volumes recorded (Figure 1), most of the