*3.4. Optimal Initial Substrate pH*

The optimization of initial pH is an important parameter to increase pigment production. Figure 4 shows the effect of initial pH (4.5, 5.5, 6.5, 7.5, and 8.5) of PP on pigment production during SSF with *A. carbonarius*. During the incubation period, the highest pigment production was determined as 61.84 ± 2.16 AU/g dfm at 400 nm for 15 d for initial pH 6.5 (Figure 4A). Moreover, 11.52 ± 1.01 AU/g dfm (Figure 4B) and 7.56 ± 1.03 AU/g dfm (Figure 4C) were measured for orange and red pigments for 15 d, respectively. It was found that as pH increased between pH 4.5–6.5, pigment production increased. However, pigment production decreased after pH 6.5. Therefore, optimum pH was determined as pH 6.5 for the pigment production at the maximum yield in this study. A similar result was obtained, so that the production of pigment by *Aspergillus nidulans* increased when the initial pH of the substrate was at 6.8 compared to pH 8.0 [56]. Furthermore, Afshari et al. [57] studied pigment production from another filamentous fungal species *Penicillium aculeatum* and they found that the best production of yellow pigment was obtained with a pH value of 6.5. The effects of pH on fungal pigment production are connected with changes in enzyme activity [56]. It is known that the optimal pH range is 5.0–6.5 for most fungi [58]. Therefore, it is thought that the growth of fungi on PP and its enzyme activity for pigment production could affect pigment production.

**Figure 3.** The effect of PP particle sizes on pigment production at (**A**) 400 nm, (**B**) 475 nm, (**C**) 500 nm during SSF with *A. carbonarius* (experimental conditions: pH: 4.5).
