## 3 Results and Discussion  
### Effect of pH on OMW Thermophilic DF  
During the optimization of OMW thermophilic DF in terms of H\({}_{2}\) and/or metabolites production, a series of batch experiments was initially performed for the pH values 4.5, 5.0, 6.0, 7.0, and 7.5 under the organic loading of 40 g COD/L. As shown in Figure 2a, only three experiments of the total five demonstrated the pH effect, and these exhibited remarkable H\({}_{2}\) yields; this was the case for the pH values of 6.0, 7.0, and 7.5, whereas the tests for the pH values of 4.5 and 5.0 indicated an insignificant production of H\({}_{2}\). The latter could be attributed to the fact that at pH values lower than 6, the enzyme hydrogenase, which is contained in H\({}_{2}\)-producing bacteria, is not able to function properly, leading to decreased or completely ceased H\({}_{2}\) production [23]. Maximum biogas (H\({}_{2}\) and CO\({}_{2}\)) production was observed for the run C (pH 6.0) at 823 mL, 45.7% of which was H\({}_{2}\). Runs D and E followed (pH 7.0 and 7.5), with H\({}_{2}\) production volumes of 205 mL and 37 mL, respectively. Convert the aforementioned values in mL H\({}_{2}\)/L\({}_{\rm reactor}\), the equivalent yields for pH 6.0, 7.0, and 7.5 are 501, 273, and 50 mL H\({}_{2}\)/L\({}_{\rm reactor}\). In terms of mol H\({}_{2}\)/mol glucose,  
Figure 4: Effect of organic loading on thermophilic OMW DF under pH 6.0.  
Figure 3: (**a**) Biogas accumulation during the test of pH 6.0; (**b**) metabolites evolution during the test of pH 6.0; (**c**) biogas accumulation during the test of pH 7.5; and (**d**) metabolites evolution during the test of pH 7.5.  
the yields followed the same pattern, with the maximum calculation of 0.70 mol H\({}_{2}\)/mol glucose for pH 6.0.  
Additionally, several main-end products such as VFAs and ethanol were measured, as described in Figure 2b. In case of runs A and B (pH 4.5 and 5.0, respectively), where significant H\({}_{2}\) production was not detected, the respective metabolites were determined in very low concentrations (less than 400 mg/L). However, for runs C, D, and E (pH 6.0, 7.0, and 7.5, respectively), the metabolites exhibited a completely different effect. Acetate was the main metabolite of the experiments, reaching a maximum concentration of 2360 mg/L at pH 7.5. For both pH values of 6.0 and 7.0, the approximate acetate concentration was 1550 mg/L. As for butyrate, its production was strongly favored at pH 6.0, reaching 942 mg/L, and then decreased in values less than 300 mg/L for pH 7.0 and 7.5. On the other hand, as pH increased (pH 7.5), the metabolites of ethanol and propionate were favored, indicating both the maximum concentrations of 955 and 802 mg/L, respectively. In the work of [PERSON] et al. [42], acetate was also the main metabolite of OMW after the DF process. However, it was followed by propionate, isovalerate, and butyrate, which could be attributed to the fact that the pH was not set to an optimum range but was left neutral (6.8-7.4), whereas the inoculum was pretreated with sodium 2-bromoethanesulfonic acid instead of heat shock. However, heat shock is still considered an efficient method for inoculum pretreatment, especially for thermophilic conditions tests [43]. Concerning the butyrate pattern versus the several tested pH values, the same trend was noticed in case of fruit and vegetable waste DF at a similar organic loading [24], indicating and ensuring that butyrate production metabolic pathway is favored in a pH range of 5.5-6.5 [44; 45].