## 2 Materials and Methods
### Cultivation  
The cultivation of the microalgae was carried out according to the previously described method [14] in a laboratory photob bioreactor (PBR) with a volume of 10 dm\({}^{3}\). Combined additives were used to stimulate the biosynthesis of the carotenoids. As a stress inducer of carotenogenesis, a 3% solution of hydrogen peroxide (\"Innovation\", Voronezh, Russia) was added to the nutrient medium. A solution of pyridoxine hydrochloride (\"BioChemica\", Moscow, Russia) with an initial concentration of 50 mg/mL was used to prevent oxidative stress. A culture medium should have the well-balanced content of macro- and microelements in order to cultivate microalgae [18]. Table 1 shows the composition of the growing medium consisting of components used to prepare the culture media A and B, with the addition of macroelements after the mixing thereof. The mother culture of _C. vulgaris BIN_ was removed from the refrigerating chamber (Melling YCD-519, Jackson, MS, USA) and introduced into the prepared PBR. The initial number of cells in the PBS was 8.0-8.5 million cells per mL. Then, the PBR was filled with the prepared culture medium. The stirring of the system during cultivation was carried out by bubbling with a Xilong AP-001 (Shatou, China) compressor at 1.5 L/min, and once a day with a mechanical stirrer for 15 min at 500 rpm (to prevent the formation of stagnant zones). The temperature range of the cultivation was 23 \(\pm\) 3 \({}^{\circ}\)C. The illumination modes--\"day-night\" (16-8 h)--were maintained using LED TIS-15M1R lamps with a luminous flux of 3000 lumens [19]. The change in the concentration of cells in the culture during the cultivation was monitored by the daily measurement of the optical density on a UV-1280 spectrophotometer (SHIMADZU, Duisburg, Germany) at a wavelength of 750 nm. The culture medium was used as a reference solution. The change in pH in the PBR was determined by using a Mini PH Meter (AQUA-LAB, Russia, Moscow). In order to study the morphological \begin{table}
\begin{tabular}{c c c c c} \hline \hline \multirow{2}{*}{**Substance**} & \multicolumn{4}{c}{_Culture Medium A_} \\ \cline{2-5}  & **Molecular Weight,** & **Concentration in Stock** & **Concentration in Culture** & \\  & **(g/mol)** & **Solution, (mg/L)** & **Medium, (ug/L)** & **Consumption, (mL/L)** \\ \hline ZnSO\({}_{4}\cdot\)7H\({}_{2}\)O & 287.53 & 0.1 & 100 & 10.0 \\ \hline CuSO\({}_{4}\cdot\)5H\({}_{2}\)O & 249.66 & 0.1 & 10 & 0.1 \\ \hline CoSO\({}_{4}\cdot\)7H\({}_{2}\)O & 281.06 & 0.1 & 100 & 5.0 \\ \hline MnCl\({}_{2}\cdot\)4H\({}_{2}\)O & 197.91 & 0.1 & 500 & 5.0 \\ \hline H\({}_{3}\)BO\({}_{3}\) & 61.83 & 0.1 & 50 & 0.5 \\ \hline Na\({}_{2}\)MoO\({}_{4}\cdot\)2H\({}_{2}\)O & 241.96 & 0.1 & 100 & 0.1 \\ \hline \multicolumn{5}{c}{_Culture Medium B (Microelements)_} \\ \hline \multirow{2}{*}{**Substance**} & \multirow{2}{*}{**Molecular Weight,** & **Concentration in Stock** & **Concentration in Culture** & \\  & **(g/mol)** & **Solution, (mg/L)** & **Medium, (ug/L)** & **Consumption, (mL/L)** \\ \hline FeCl\({}_{3}\cdot\)6H\({}_{2}\)O & 270.21 & 1.0 & 4.000 & 4.0 \\ \hline Na\({}_{2}\)EDTA\(\cdot\)2H\({}_{2}\)O & 372.24 & 1.0 & 6.000 & 6.0 \\ \hline \multicolumn{5}{c}{_Macroelements_} \\ \hline \multirow{2}{*}{**Substance**} & \multirow{2}{*}{**Molecular Weight,** & **Concentration in Mother** & \\  & **(g/mol)** & **Medium, (mg/L)** & **Consumption, (g/L)** \\ \hline KNO\({}_{3}\) & 101.1 & 1.000 & 3.26 \\ \hline KH\({}_{2}\)PO\({}_{4}\) & 136.07 & 100 & 0.32 \\ \hline MgSO\({}_{4}\cdot\)7H\({}_{2}\)O & 246.48 & 240 & 2.46 \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Composition of the microalgae culture medium. characteristics of the _C. vulgaris_ cells, digital images of the \"crushed drop\" preparations were used with a magnification of at least 400 times using an IS-500 digital camera (LOMO JSC, St-Petersburg, Russia). The digital images were processed using the FOTO Microanalysis software (LOMO JSC, St-Petersburg, Russia) and Levenhuk (Levenhuk LabZZ manufacturer, China).