## 2 Materials and Methods
### Experimental Setup  
The experiment was carried out in vertical tubular photobioreactors (Aqua Medic, Bissendorf, Germany) with a capacity of 2.5 \(\mathrm{d}\mathrm{m}^{3}\) (Figure 1), supplemented with 2 \(\mathrm{d}\mathrm{m}^{3}\) of the F/2 culture medium [19] with the composition (g/L): NaNO3--0.075 g; NaH2 PO4 2H2O --0.00565 g; stock solution of trace elements--1 mL/L (Na2 EDTA 4.16 g, FeCl3 6H2O 3.15 g, CuSO4 5H2O 0.01 g, ZnSO4 7H2O 0.022 g, CoCl2 6H2O 0.01 g, MnCl2 4H2O 0.018 g, and NaMoO4 2H2O 0.018 g) and stock solution of vitamin mix--1 mL/L (cyanocobalamin (vitamin B12) 0.0005 g, thiamine HCl (vitamin B1) 0.1 g, biotin 0.0005 g). After sterilization of the medium with UV light, 200 \(\mathrm{c}\mathrm{m}^{3}\) of C. _volaris_ microalgae inoculum was introduced into the photobioreactors. In the study, we analyzed the effect of two types of lighting: high-pressure sodium light (HPS-Son-T-Agro, 400 watt, light intensity of 660 \(\mathrm{\SIUnitSymbolMicro mol\cdot s^{-1}}\)/56,000 lux; PHILIPS, Amsterdam, Netherlands) and light-emitting diode (LED light with white diodes, red diodes (wavelength 600-700 nm, light intensity of 9.45 \(\mathrm{\SIUnitSymbolMicro mol\cdot s^{-1}}\)) and blue diodes (wavelength 400-500 nm, light intensity of 2.25 \(\mathrm{\SIUnitSymbolMicro mol\cdot s^{-1}}\)). Total LED light intensity was 13.5 \(\mathrm{\SIUnitSymbolMicro mol\cdot s^{-1}}\) (HOLDBOX, Zabia Wola, Poland). Three times of lighting culture were used: 12/12, 18/6, and 24/0 h in the light/dark cycle and four pH levels were used: 6, 7, 8, and 9. Algae cells were kept suspended by mixing with gas using a 25 W pump (Aqua Medic, Bissendorf, Germany) with a capacity of 0.9 \(\mathrm{m}^{-3}\mathrm{.h}^{-1}\). At the same time, the aeration made it possible to introduce carbon dioxide into the culture. The experiment was carried out as a batch culture. To establish an adequately fast and accurate method for the quantification of microalgae biomass, three comparative methods were used: biomass dry weight, measuring the content of chlorophyll a, and counting the number of cells using a hemocytometer. The dry weight of microalgae has been determined by the gravimetric method described by [PERSON] et al. [20] with modification. Biomass estimation was carried out using a moisture analyzer (AXIS ATS, Gdarask, Poland). Aliquots of 40 cm\({}^{3}\) were centrifuged (Eppendorf, Hamburg, Germany) at 4000\(\times\)\(g\) for 15 min. The biomass after centrifugation was transferred to aluminum plates, dried at 105 \({}^{\circ}\)C to constant weight, cooled, and weighed. The results were calculated and presented in mg\(\cdot\)L\({}^{-1}\). The amount of biomass was determined on the day the culture was established and at 24-h intervals for the next 15 days. The biomass productivity was calculated by the following equation  
\[\text{BP}=\frac{\text{B}_{\text{f}}-\text{B}_{0}}{\text{d}},\]  
where BP is the biomass productivity (mg\(\cdot\)L\({}^{-1}\cdot\)d\({}^{-1}\)), \(\text{B}_{\text{f}}\) and \(\text{B}_{0}\) are final and initial biomass concentration (mg), respectively, and d is the cultivation time (day). Chlorophyll concentration was determined according to the PN-86 C-05560/02 standard [21]. The pigment was extracted from biomass concentrated by filtration on a glass fiber filter using 90% (\(v/v\)) acetone. The absorbance of acetone extract was measured at wavelength of 665 nm, with a spectrophotometer (EMCO, Warszawa, Poland). The results are given in mg m\({}^{-3}\). The chlorophyll productivity was calculated using following equation:  
\[\text{ChP}=\frac{\text{Ch}_{\text{f}}-\text{Ch}_{0}}{\text{d}},\]  
Figure 1: Vertical tubular photobioreactors used in the research. where ChP is chlorophyll productivity (mg:m\({}^{-3}\).d\({}^{-1}\)), Ch\({}_{\text{f}}\) and Ch\({}_{0}\) are final and initial content of chlorophyll (mg), respectively, and d is the cultivation time (day). Microalgal cells were counted using Thoma hemocytometer [22]. Two chambers of the hemocytometer were filled up with culture samples.