# Application of Extracellular Polymeric Substances during Cultivation of Microalgae Biomass
## 3 Results and Discussion
\begin{table}
\begin{tabular}{c c c c c c} \hline  & **Variant of** & **Biomass** & **Chlorophyll** & **EPS Content** & **Sedimentation** \\  & **Experiment** & **Concentration (mg/L)** & **Content (mg/g VS)** & **(mg TOC/g VS)** & **Efficiency (\%)** \\ \hline  & C & 3621 \(\pm\) 174 & 40 \(\pm\) 11 & 95 \(\pm\) 11 & 40 \\ _Tetraselmis_ & V1 & 4109 \(\pm\) 160 & 11 \(\pm\) 4 & 97 \(\pm\) 7 & 42 \\ _subcordiformis_ & V2 & 3698 \(\pm\) 102 & 32 \(\pm\) 9 & 97 \(\pm\) 25 & 55 \\  & V3 & 3856 \(\pm\) 111 & 28 \(\pm\) 12 & 97 \(\pm\) 10 & 66 \\ \hline  & C & 3920 \(\pm\) 289 & 28 \(\pm\) 8 & 60 \(\pm\) 20 & 32 \\ _Chorella_ sp. & V1 & 4210 \(\pm\) 320 & 8 \(\pm\) 6 & 61 \(\pm\) 9 & 31 \\  & V2 & 4290 \(\pm\) 255 & 13 \(\pm\) 5 & 62 \(\pm\) 12 & 34 \\  & V3 & 4321 \(\pm\) 262 & 10 \(\pm\) 2 & 61 \(\pm\) 19 & 36 \\ \hline  & C & 1213 \(\pm\) 182 & 16 \(\pm\) 4 & 81 \(\pm\) 6 & 49 \\ _Arthrospien platensis_ & V1 & 1512 \(\pm\) 191 & 8 \(\pm\) 4 & 83 \(\pm\) 9 & 54 \\  & V2 & 1760 \(\pm\) 195 & 11 \(\pm\) 6 & 91 \(\pm\) 12 & 68 \\  & V3 & 1912 \(\pm\) 150 & 9 \(\pm\) 4 & 96 \(\pm\) 2 & 72 \\ \hline \end{tabular}
\end{table} 
Table 1: Biomass concentrations, chlorophyll content, EPS content, and sedimentation efficiency obtained in the variants of experiments during the cultivation of _T. subcordiformis_, _Chlorella_ sp., and _A. platensis_. The biomass growth in the control and until 10 th day of microalgae/cyanobacteria cultivation in V2 and V3 was the same since on the 10 th day after EPS were added to the control cultivation. The biomass concentration during strategy V1 was higher than during the control for all tested microalgae/cyanobacteria. During V1, the biomass concentration of _T. subcondiformis_ and _Chlorella_ sp. was similar, and the growth rate was around 600 mg/(L-d) in the first 5 days of cultivation and around 100 mg/(L-d) in the next 5 days (Figure 2). The _A. platensis_ growth rate was steady during cultivation, being around 150 mg/(L-d). Figure 2: _Cont._  
Microalgae growth generally can be divided into six phases during batch cultivation: the lag phase, exponential phase, linear phase, declining growth phase, stationary phase, and death phase. In the lag phase, the growth of the cell is delayed due to the presence of non-viable cells or adaptation to new environmental conditions (the addition of organic carbon). In the present study, the lag phase was not observed for all tested strategies. Similar observations for _Chlorella vulgaris_ were noted during transcriptomic studies of the transition of photoautotrophic to mixotrophic metabolism, in which biomass after the addition of glucose started an exponential phase [22]. Additionally, the studies of terrestrial cyanobacteria cells under heterotrophic and mixotrophic conditions did not show a lag phase [28]. The lag phase for _C. sorokiiuna_ during mixotrophic cultivation with different volatile fatty acids lasted less than 1 day, which can also be the explanation for observations in the present study because the measurements were taken once a day [29]. The above-mentioned explanations support the statement of mixotrophic metabolism observed during EPS addition in strategy V1. The mixotrophic cultivation did not increase the sedimentation efficiency of _T. subcoriformis_ and _Chlorella_ sp. biomass (Table 1). The biomass of _A. platensis_ was characterized by a 5% better efficiency of sedimentation after mixotrophic cultivation than autotrophic cultivation. The introduction of EPS at the end of cultivation (V2 and V3) also lowered chlorophyll content in biomass; however, the reduction was not significant (Table 1). In V3, the chlorophyll content was lower than in V2. The higher concentrations of source of organic carbon available for biomass, the greater the reduction in the chlorophyll content in biomass. In V2 and V3, the chlorophyll content was reduced by about 25%, 60%, and 38% compared to autotrophic cultivation in the biomass of _T. subcoriformis_, _Chlorella_ sp., and _A. platensis_, respectively.