## 3 Results and Discussion
### Two-Stage Anaerobic Digestion of Wheat Straw and Algal Biomass
#### 3.2.1 Influence of HRT  
Changing the HRT led to variations in the energy carrier yields (Figures 4 and 5).  
The maximum daily concentration of hydrogen was achieved at HRT of 2 days, at 42.5% H\({}_{2}\), and the maximum concentration of methane reached was 56.1% at an HRT of 6 days.  
The results of the average concentration are comparable to those of the maximum yield, the best being the average yield of hydrogen at an HRT of 2 days--120.9 mL/day/g VS\({}_{\text{add}}\)--and correspondingly, the best average yield of methane was 581.8 mL/day/g VS\({}_{\text{add}}\) at HRT = 6 days.  
The HRT is a very important parameter during hydrogen and methane production [25]. Short HRTs are associated with hydrogen fermentation. Hydrogen production occurs at lower retention times and depends on the feed flow rate. The hydrogen-producing bacteria  
Figure 4: Hydrogen yield in the two-phase anaerobic digestion system depending on the HRT in the first bioreactor.  
Figure 5: Methane yield in the two-phase anaerobic digestion system depending on the HRT time in the first bioreactor.  
that reside in BR-1 are fast growing, but short HRTs or high dilution rates could be applied to eliminate methanogens, which have extremely low growth rates [26]. In BR-2, the HRT describes the average times that certain substrates reside in a digester, which are higher in this reactor. In this case, if the HRT is shorter, the system could stop due to washout of microorganisms.  
Adjusting the appropriate operational parameters of bioreactors, such as increasing the HRTs and organic loading rates, is a way to increase biodegradation efficiencies [27]. The recalcitrant nature of lignocellulosic biomass hinders its efficient exploitation for energy production [28]. However, lignocellulosic has been exploited as a substrate for biogas production due to its high energy potential. The cellulose concentration was determined to deplete during the process (2.75% in the methanogenic bioreactor at the end of the process).  
The H\({}_{2}\)S concentration in biogas varied from 100 to 10,000 ppmv depending on the feedstock's sulphur content. Our measurements showed values from 200 to 283 ppmv. No methanogenesis inhibition was registered. There is also a threshold concentration of H\({}_{2}\)S in biogas which will impact further applications, so technologies are being developed for H\({}_{2}\)S removal from biogas [29].  
Utilization of algal biomass for the production of gaseous biofuels using methane as an energy carrier supports a circular economy. In the digestion of algae, all cell compounds--sugars, fats, nucleic acids and proteins--could be involved. While protein in algal cells can result in undesirable ammonia formation, other industrial waste that is rich in carbon but poor in nitrogen compounds, like wheat straw, could be added [30]. The moisture and nutrient content of microalgae can favour lignocellulosic waste digestion.  
Anaerobic digestion of algal biomass implies costs reductions via the elimination of drying wet biomass and extraction before processing, as they become unnecessary [31]. The increase in cellulase activity might be favourable for the biodegradation of algal biomass and straw, which could supply the bioreactors with nutrients and finally increase the methane production rate.