# Propagation of Inoculum for _Haematococcus pluvialis_
## 2 Materials and Methods  
The _Haematococcus pluvialis_ G1002 microalgae strain was used to conduct the experiment. This strain comes from the CAUP culture collection (Charles University in Prague, Czech Republic). The tests were conducted using the Bold's Basal Medium (BBM), which was sterilized at 121\({}^{\circ}\)C, for 15 min, in the Tuttnauer 5075 ELV-D WR laboratory autoclave (Breda, Netherlands). The microalgae cultivation was conducted over 8 days in the laboratory room, in a temperature range of 23-25\({}^{\circ}\)C. The pH value was maintained automatically at 7.00 (-) and with hysteresis at the level of 0.2 (-). It was regulated by introducing carbon dioxide into the photobioreactors. Over the course of the experiments, a manual setting of the intensity of the LED lighting was used at the level of 54% of power throughout the course of cultivation. Building the workstation required using eight glass air-lift bioreactors with a total volume of 12 dm\({}^{3}\) each, which were designed by AlgaeLabs (Wroclaw, Poland). The photobioreactors were built out of two glass tubes. The walls of the tubes were 5 mm thick (Schott). An internal tube (riser), which is 125 mm in diameter and 400 mm in length, was placed inside an external pipe (downcomer), which is 190 mm in diameter and 500 mm in length. Thus, obtained photobioreactors have a total volume of 11.97 dm\({}^{3}\). The photobioreactors were covered with a top dome, which included necessary ports for connecting the temperature, and pH sensors. The bottom dome was fitted with an enclosed diffuser that allowed for the effective atomization of the introduced gases and the nozzle responsible for filling and emptying the photobioreactor. The photobioreactors were sterilized using a 4% solution of perhydrol. The photobioreactors were filled with perhydrol for a period of two hours. Subsequently, they were rinsed with deionized water. Bioreactors, which were prepared in such manner were filled with culture medium and the microalgae inoculant and supplemented with deionized water to obtain 10 dm\({}^{3}\) of working volume. Photobioreactors temperature measurements were conducted using thermo-resistant sensors STPt 100 (Hydromet, Gliwice, Poland) in a borosilicate glass vessel, which is resistant to aggressive solutions such as concentrated alkalis, concentrated and diluted acids (including oxidizing acids) as well as organic-water mixtures. The sensors were capable of measuring the temperature within the 0 and 100 \({}^{\circ}\)C range. The sensors cooperated with temperature transmitters with the RS485 interface and Modbus RTU (AlgaeLabs, Wroclaw, Poland) communication protocol. Composite electrode ERH-13X2(Hydromet, Gliwice, Poland) in a glass case was used to measure pH. This electrode was intended for measuring the pH in water or water solutions in industrial applications. The pH sensors cooperated with pH measurement transducer with the RS485 interface, and Modbus RTU (AlgaeLabs, Wroclaw, Poland) communication protocol. The lighting of the photobioreactors was designed using EMPA-d (epiLED, Wroclaw, Poland) type strips with electroluminescent LED diodes, powered by 12 V DC, color temperature- warm white 2800-3200 K, with a 120\({}^{\circ}\) beam angle and light output of 9600 lumens. Power consumption of 96W/5 m of the LED strip with 120 diodes per 1 m of LED strip, type 2835 (REFOND Optoelectronics, Shenzhen, China) were located on 1 m of LED strip. Four light panels with 0.5 m long LED strips were prepared for each station. Diodes and strips drivers LED RGB SPL-3C (Dagon, Leszno, Poland) were used to control the LED lighting. The source of the voltage signals 0-10 V was an 8-channel analog output module SDM-8 AO (SFAR) with serial interface RS-485 and Modbus RTU communication protocol. Electrically controlled solenoid valves 322 ME (AZ Pneumatica, Misinto, Italy) type 3/2 NC with 24 V DC coils were used to control the gas flow.