Effect of Photoperiod and Glycerol Supplementation on the Biomass Productivity and Protein Production of Spirulina sp. LEB 18 Cultures
, Itaciara Larroza Nunes 2, Janice Izabel Druzian 3, Michele Greque de Morais 4, Ana Priscila Centeno da Rosa 1 and Jorge Alberto Vieira Costa 1,*
Round 1
Reviewer 1 Report
The manuscript entitled Effect of photoperiod and glycerol supplementation on the biomass productivity and protein production of Spirulina sp. LEB 18 cultures, present information related to the production of biomass, protein, chlorophyll, and fatty acids by Spirulina sp. LEB 18. The manuscript presents issues that authors must attend to prior to acceptance.
-A native English speaker must revise the language.
-Introduction must provide detailed information on the importance of producing biomass, protein, and fatty acids. Basically, justify why authors developed the experiments and what they solve with the results obtained.
-Lines 56 and 58. Spirulina must be written in italics. Revise the same issue in the manuscript.
-Line 64. Authors must identify the microalgae at the species level.
- Line 72. Inform the used aeration flow rate.
-Line 85. How was determined the logarithmic phase?
Line 146. How was defined the lag phase?
Lines 148-149. How authors concluded the control treatments did not show an exponential phase? Did the authors model the data?
Author Response
Response to Reviewer 1 Comments
Point 1: A native English speaker must revise the language
Response 1: The manuscript was revised for English, as suggested.
Point 2: Introduction must provide detailed information on the importance of producing biomass, protein, and fatty acids. Basically, justify why authors developed the experiments and what they solve with the results obtained.
Response 2: Thank you for the suggestion, we changed our introduction on lines 54-58, explaining the importance of high biomass productivity on microalgae cultures, and from lines 60-65, we justify the importance of the research of Spirulina strain and its protein and fatty acids content.
Point 3: Lines 56 and 58. Spirulina must be written in italics. Revise the same issue in the manuscript.
Response 3: Thank you very much for this observation. We review and correct all microorganism names to italics in the manuscript.
Point 4: Line 64. Authors must identify the microalgae at the species level.
Response 4: Our Spirulina sp LEB 18 strain is not identified to the species level, and we can’t do the genomic analysis at our lab in time for this article. However, our group has widely used this strain since its isolation in Morais et al. (2008); we added this reference to the manuscript (line 74). Several publications on this specific strain application can be found in the literature. We will consider your suggestion for future publication with this strain.
Point 5: Line 72. Inform the used aeration flow rate.
Response 5: The aeration flow rate used was 0.5 L min-1. We add this to the text (line 83) and reference previous studies that use the same condition (MORAIS et al., 2022a).
Point 6: Line 85. How was determined the logarithmic phase?
Response 6: The logarithmic/exponential phase occurs when the biomass x time graph curve shows exponential growth, and the specific growth rate is constant and maximum. To identify the logarithmic phase of microalgal growth, we build a curve with the log of biomass concentration (g L-1) vs time (h) using excel. In this graph, the logarithmic phase appears as a straight line that, with a linear regression, shows an R2 close to 1, and with this, we can obtain the line equation (y=ax+b).
Point 7: Line 146. How was defined the lag phase?
Response 7: According to Posten and Cooney (2001), the lag phase is defined as the phase in which organisms do not significantly increase in number but are metabolically active: they grow, synthesize enzymes and incorporate nutrients present in the medium. Therefore, the lag phase is the adaptation of microorganisms to the environment, in which the biomass concentration does not increase over time and precedes the log phase.
Point 8: Lines 148-149. How authors concluded the control treatments did not show an exponential phase? Did the authors model the data?
Response 8: We conclude that the control treatments did not show an exponential phase based on the observation of the growth curve. On 36h of culture, cells don’t show exponential growth as the cell concentration remains constant, differently from the cultures supplemented with glycerol that show exponential growth and a constant specific growth rate.
References
MORAIS, E. G. DE; NUNES, I. L.; DRUZIAN, J. I.; DE MORAIS, M. G.; DA ROSA, A. P. C.; COSTA, J. A. V. Increase in biomass productivity and protein content of Spirulina sp. LEB 18 (Arthrospira) cultivated with crude glycerol. Biomass Conversion and Biorefinery, v. 12, n. 3, p. 597–605, 2022a.
MORAIS, E. G. DE; NUNES, I. L.; DRUZIAN, J. I.; MORAIS, M. G. DE; ROSA, A. P. C. DA; COSTA, J. A. V. Increasing the cell productivity of mixotrophic growth of Spirulina sp . LEB 18 with crude glycerol. Biomass Conversion and Biorefinery, n. 0123456789, p. 1–9, 2022b.
MORAIS, M. G. DE; REICHERT, C. DA C.; DALCANTON, F.; DURANTE, A. J.; MARINS, L. F.; VIEIRA COSTA, J. A. Isolation and Characterization of a New Arthrospira Strain. Zeitschrift für Naturforschung C, v. 63, n. 1–2, p. 144–150, 2008.
POSTEN, C. H.; COONEY, C. L. Growth of Microorganisms. In: Biotechnology Set. [s.l.] Wiley, 2001. p. 111–162.
Author Response File: 
Author Response.docx
Reviewer 2 Report
This manuscript is devoted to the study of the effect of glycerin on biomass productivity and the production of Spirulina sp. LEB18 cultures protein at various photoperiods. The authors conducted the necessary experiments and performed their processing and interpretation.
There are the following questions and recommendations:
1. Which LED lamps were used for experiments? What is their radiation spectrum?
2. Why was the irradiance chosen exactly 60 µmolfótons.m-2 s-1 ?How was it controlled during the experiment?
3. Figures 1 and 2 should be colored.
Author Response
Response to Reviewer 2 Comments
Point 1: Which LED lamps were used for experiments? What is their radiation spectrum?
Response 1: The LED lamps used is High Flux LED Bulb HO OSRAM 40W (white light) and was identify. The radiation spectrum is of the white light (400 – 700 nm). Information was added on lines 79 to 81
Point 2: Why was the irradiance chosen exactly 60 µmolfótons.m-2 s-1? How was it controlled during the experiment?
Response 2: The luminosity was monitored by a digital radiometer (WALZ ULM-500, Germany) at the beginning and at the end of the experiment; both showed a constant irradiation of 60 µmolfótons.m-2 s-1, this information were add at line 80. This condition was chosen because it was applied by the literature like in the article published by MORAIS et al. (2022).
Point 3: Figures 1 and 2 should be colored.
Response 3: The figures were colored as suggested by the reviewer.
References
MORAIS, E. G. DE; NUNES, I. L.; DRUZIAN, J. I.; MORAIS, M. G. DE; ROSA, A. P. C. DA; COSTA, J. A. V. Increasing the cell productivity of mixotrophic growth of Spirulina sp . LEB 18 with crude glycerol. Biomass Conversion and Biorefinery, n. 0123456789, p. 1–9, 2022.
Author Response File: Author Response.docx
Round 2
Reviewer 1 Report
The manuscript has been improved. I recommend accepting in the current form.
