## 3 Results and Discussion
### Tolerance of the Microalgae to Raw Whey (RW) Wastewater  
The tolerance of both _Chlorella_ sp. and _Scenedesmus_ sp. to RW diluted with distilled water in different proportions (\(v/v\)) was explored in batch flask cultures (Section 2.3), coded as follows: RW-10%, RW-20%, RW-30%, RW-40%, and RW-50%. Figure 3 shows semilog plots illustrating the normalized cell concentration for the surviving microalgae treatments (RW-10%, 20%, and 30%): Figure 3A for _Chlorella_ sp. and Figure 3B for _Scenedesmus_ sp. Inaddition, results from the effluents generated during the pretreatment of the different RW (coded as TRW-10%, TRW-20%, and TRW-30% in Section 2.4) in PBR-1 for the sequential batch photobioreactor experimentation, which were also tested for tolerance in flask cultures, are included in Section 3.3.2 for later comparison. No apparent lag phase was observed for any of them, regardless of the cultured species. The onset of the exponential growth phase occurred at the beginning of cultivation and extended until around the third day for both microalgae. Throughout this phase, the maximum specific growth rates were determined through regression analysis using Equation (3). Except for the BG11 control, the RW-based cultures did not have a linear phase, and therefore their exponential phase was followed by a stationary phase. Based on the growth kinetics depicted in Figure 3A,B, \(\mu_{max}\), Y\({}_{B}\), and \(P_{B}\) were calculated as detailed in Section 2.5. A two-way ANOVA was performed to assess the influence of RW% and microalgal species (ALGA) on each of the three response variables. The results revealed that both RW% and ALGA had a statistically significant effect (\(p<0.001\)) on: (i) \(\mu_{max}\) (F (3, 16) = 30.4), F (1, 16) = 28.3, respectively); (ii) \(Y_{B}\) (F (3, 16) = 101.5), F (1, 16) = 28.1, respectively); and (iii) \(P_{B}\) (F (3, 16) = 26.9), F (1, 16) = 24.2, respectively). Moreover, a significant interaction (\(p<0.001\)) between RW% and ALGA was observed (\(\mu_{max}\), F (3, 16) = 9.5; Y\({}_{B}\), F (3, 16) = 7.6; \(P_{B}\), F (3, 16) = 8.75), indicating that the combined effects of these factors were not independent. According to the F values, the percentage of the variance was distributed from highest to lowest in the following order: RW%, ALGA, and interaction. Figure 3D-L presents the outcomes of the two-way ANOVA. Only RW-10% significantly improved the mean \(\mu_{max}\) (0.835 d\({}^{-1}\)) compared to the control group (BG11 medium) (Figure 3D). Although no statistically significant differences were observed between the RW-20% and RW-30% groups relative to the control group, their values were lower than RW-10%, indicating an inhibitory effect due to excess substrate; the higher the RW%, the lower the \(\mu_{max}\). The group of _Chloerella_ sp. MC18 (CH) cultures exhibited a mean \(\mu_{max}\) value, 0.760 d\({}^{-1}\), 1.12-fold higher than that of _Scenedensmus_ sp. MJ23-R (SC) (Figure 3E). Figure 3F highlights that the effect of the RW% on \(\mu_{max}\) differed within the group ALGAE. The increase in _Chloerella_\(\mu_{max}\) at RW-10% relative to the control was more pronounced compared to _Scenedensmus_. The RW% and ALGAE factors affected in a similar way to \(P_{B}\) (Figure 3G-I). Conversely, the mean \(Y_{B}\) exhibited a downward trend with a higher RW% relative to the control group (Figure 3J), suggesting potential nutrient imbalance, toxicity, stress responses, or metabolic burden. While these hypotheses warrant further experimental validation, they offer plausible explanations for the observed trend. Future research efforts could focus on elucidating these mechanisms to enhance our understanding of microalgal growth dynamics in RW-based cultures. The mean \(Y_{B}\) of the _Chloerella_ culture group was higher than that of _Scenedensmus_ as observed for \(\mu_{max}\) and \(P_{B}\) (Figure 3K). However, the interaction between RW and ALGA was manifested for the RW-20% and RW-30% groups, for which \(Y_{B}\) was higher in _Chloerella_ than in _Scenedensmus_ (Figure 3L).