# The Effect of Hydrolysis on the Antioxidant Activity of Olive Mill Waste
## 4 Discussion
### Antioxidant Activity
#### 4.2.1 Radical Scavenging Activity  
_DPPH Radical Scavenging_ activity preferably reacts via SPLET mechanism in solvents such as methanol and ethanol; this is because it tends to be more stable [77]. From the results obtained during this study, it was observed that alkaline hydrolysis had the best scavenging activity compared to acid hydrolysis and methanolic macerated extracts. This was seen in the study conducted by [PERSON], [PERSON] and [PERSON] [78], where it was stated that non-hydrolysed extracts had a lower antioxidant activity, whilst acid and alkaline hydrolysis had a higher antioxidant activity. In comparison to the results obtained, [PERSON] et al. [38] found that the scavenging activity was higher in acidic hydrolysis compared to non-hydrolysed and alkaline hydrolysis.  
_ABTS_ assay preferably reacts via the SPLET mechanism in aqueous solutions [77]. From the results obtained, _ABTS_ radical cation stabilization had a better scavenging activity compared with _DPPH_. This could be because the _ABTS_ assay tends to be more sensitive for identifying antioxidant activity, since it has faster reaction kinetics and a heightened response to antioxidants [79]. In contrast, the results obtained by [PERSON] et al. [80] state that the IC\({}_{50}\) values for the _DPPH_ scavenging assays of all extracts were slightly higher than those of _ABTS\({}^{\bullet+}\)_. In this study, it was noted that acidic hydrolys had the lowest IC\({}_{50}\)'s compared with alkaline and methanolic extracts. Furthermore, [PERSON] et al. [38] showed that the highest scavenging activity was seen in alkaline hydrolysis rather than in acidic hydrolysis, with a scavenging activity of 96%. Similar results were seen in the study on the effect of hydrolysis conditions on recovery of antioxidants from methanolic extracts of Nigella Sativa seeds conducted by [PERSON], [PERSON] and [PERSON] [78], where it was stated that alkaline hydrolysis had the highest antioxidant activity.  
From the correlations obtained in Table 4 for _DPPH_ and _ABTS_ with TPC, TFC and TdOPC correlated best with methanolic extracts. This is in line with studies by both [PERSON] et al. [81] and [PERSON] et al. [82], where it was stated thatTPC strongly correlated with _DPPH_ in methanolic extracts. Another study by [PERSON] et al. [38] states that the highest TFC was found in acidic hydrolysis rather than in neutral or alkaline hydrolysed extracts. In this study, it was stated that TPC correlated with _DPPH_ and _ABTS_, but there was no correlation between the three extraction procedures used, including a non-hydrolysed, basic and acidic hydrolysis extraction. The correlations for _DPPH_ and _ABTS_ with TPC, TFC and TdOPC tend to be much less for acid and alkaline hydrolysed extracts than for methanolic extracts. This could be because compounds are being released during hydrolysis; thus, an increase in antioxidant activity is seen, but a weaker correlation can be observed. In 2015, [PERSON], [PERSON] and [PERSON] [83] conducted a study on the reactivity of phenolic compounds towards free radicals under in vitro conditions. In this study, it was concluded that hydroxylated cinnamates are more effective than their benzoic acid counterparts. This could be due to the bulky -CH=CH-COOH group, which increases the activity by stabilizing the resulting phenoxy radicals [84]. Furthermore, the double bond serves as a way to stabilize the radicals of the cinnamic acid derivatives via resonance [85; 86]. Thus, comparing the correlations observed with the literature, methanol extracts tend to contain more complex molecules which show a better correlation with antioxidant activity due to the presence of the conjugated compounds. The presence of the unhydrolyzed glucoside groups has shown that they might enhance their stability in the presence of glucose moieties that decrease the antioxidant activity. This is because glycosides increase steric hindrance during the reaction [87].