Outdoor Large-Scale Cultivation of the Acidophilic Microalga Coccomyxa onubensis in a Vertical Close Photobioreactor for Lutein Production
, Mari-Carmen Ruiz-Domínguez 2, Benito Mogedas 1, Inés Garbayo Nores 1,*, Manuel González del Valle 3 and Carlos Vílchez 1Round 1
Reviewer 1 Report
Dear Author
The work presented is very interesting. The presented method of biomass cultivation in microalgae is a new element.
However, the work requires corrections.
Line 57-58. You write about the antibacterial effect of microalgae. Please edit in detail what antagonism mechanisms are observed. (maximum 10 sentences).
Line 77-85: Please unify the units specifying the composition of the substrate.
Line 142-146: Please provide the conditions for determining lutein: column, mobile phase, mobile phase flow rate.
Table 1.: In table 1 please first give the results of your experiments and then literature data.
Please perform a statistical analysis of your experiments. Perhaps the breeding methods have an impact on the efficiency of lutein synthesis.
In conclusion, please specify the conclusions clearly.
Author Response
We thank the reviewer for the constructive comments which allowed us to improve our manuscript. Please see below our answers to his/her comments.
REVIEWER 1.
Comment 1. - Line 57-58. You write about the antibacterial effect of microalgae. Please edit in detail what antagonism mechanisms are observed. (maximum 10 sentences).
Answer. The following paragraph has been included (from line 56): “Among the potential applications of this microalga, antibacterial effect against human pathogens has been reported by Navarro et al. [13], and microalgal fatty acids were suggested to be involved in the antibacterial activity. Although the mechanisms through which fatty acids may exert bactericidal activity are not fully understood, it has been reported they may promote membrane damage resulting in nutrients uptake alteration and cellular respiration inhibition [14].”
Comment 2.- Line 77-85: Please unify the units specifying the composition of the substrate.
Answer: It has been corrected.
Comment 3.- Line 142-146: Please provide the conditions for determining lutein: column, mobile phase, mobile phase flow rate.
Answer: The following paragraph has been included (from line 152): “Specific carotenoids—lutein and β-carotene—were separated by HPLC equipped with a diode-array detector (L-7420, TermoQuest, CA, USA) and a RP18 column (LichroCart RP18, Merck KGaA, Darmstadt, Germany), 5µm, size 250 x 4 mm. In the mobile phase, solvent A was ethyl acetate and solvent B was acetonitrile and water (9:1, v/v) (mobile phase flow rate was 1 mL per minute). Carotenoids detection was at 450 nm and the carotenoids were quantified using lutein and β-carotene standards supplied by DHI-Water and Environment (Hørsholm, Denmark).”
Comment 4.- Table 1: In table 1 please first give the results of your experiments and then literature data.
Answer: Results from these experiments were first given in the table.
Comment 5.- Please perform a statistical analysis of your experiments. Perhaps the breeding methods have an impact on the efficiency of lutein synthesis.
Answer: Indeed, the volumetric productivity obtained is still low compared to other lutein accumulating microalgae (Table 1 and text, this paper). In this sense, cell density in the reactor and light irradiance, among other factors, might have a significant impact both in intracellular accumulation and volumetric productivity of lutein. The following paragraph has been included in page 8: “the cultivation system might influence the lutein accumulation in the cells, as light path varies largely depending on the production system, for instance tubular photobioreactors light path is usually shorter than that in raceway open ponds [27]. In microalgae lutein may accumulate under relatively low irradiance levels as, for instance, proven in Chlorella [28], which suggests that the low volumetric productivity shown in this paper might still be enhanced in dense cultures”.
The results shown in this manuscript correspond to a first attempt for production of Coccomyxa onubensis outdoors. In order to perform a statistical analysis, additional cultivation trials outdoors with the acidophilic microalga have to be done. To the best of our knowledge, this paper shows the cultivation attempt of an acidophilic microalga outdoors, thus exploring the potential of this type of extremophilic microorganisms for massive production.
Comment 6.- In conclusion, please specify the conclusions clearly.
Answer: “The capacity of an acidic extreme environment microalga, C. onubensis, for massive production in outdoor photobioreactors has been evaluated. The production in acidic medium minimizes the risk of biological contamination and reduces the loss of algal productivity associated with microbial proliferation. C. onubensis can be produced massively in outdoor tubular photobioreactors, at acidic pH values, during the springtime in southern Europe, seemingly with limited photoinhibition if the production is carried out at suitable biomass concentrations. Non-optimized, maximal biomass and lutein productivities of 0.14 g L–1 d–1 and 1.4 mg L–1 d–1, respectively, were achieved, which can still be further improved. The optimization of process engineering at extreme acidic pH, including pH-control and the use of acid-resistant materials, is a key challenge to maximize the productivity of C. onubensis and their derived products in photobioreactors.”.
Reviewer 2 Report
In this paper, the authors investigate outdoor large-scale cultivation of the acidophilic microalga Coccomyxa onubensis in a vertical close photobioreactor for lutein production.
The paper and the topic are interesting, but however, the following issues should be addressed before considering publication in Processes journal.
- title… Name of the microalga in title should be in italic
- row 102 … what does the temperature “below 10-25°C˝ mean? Is it between 10-25°C?
- row 115 – supplier, town and country are missing for pH meter and temperature sensor.
- Since the experiment was performed in the 2 specific months (when talking about temperature and light) can the authors provide more details about temperature control (especially heating) of the device. And what about light source when light is limited.
- Figure 2 – I would suggest putting same scale (from 0-1.8) on figures A and C for better visualisation and to correct x-axe on figure C so zero of both axes intersect
- Conclusion is missing
- There are no newer references ( 5 are from 2016 y and the rest are older) so please provide newer.
- Please check English language.
- There are many small errors in the manuscript i.e.
- row 122 …. dot at the end of the sentence is missing
- row 145 …. there is one excess bracket in the sentence
- rows 182 – 186 … some units in the paper are written with the · between them and most of them without so authors must decide what type they will use
- row 257…. C. should be in italic etc.
Author Response
We have uploaded the revised version of the manuscript, which includes changes according to the suggestions of both reviewers. All comments and suggestions have been addressed. The new changes made in the text are in track changes mode in the article word file.
We thank the reviewer for the constructive comments which allowed us to improve our manuscript. Please see below our answers to his/her comments.
Best regards,
Prof. I Garbayo
Response to Reviewer 2 Comments
REVIEWER 2.
Comment 1.- Title… Name of the microalga in title should be in italic
Answer: It has been corrected.
Comment 2.- Row 102… what does the temperature “below 10-25°C˝ mean? Is it between 10-25°C?
Answer: It has been corrected.
Comment 3.- Row 115 – supplier, town and country are missing for pH meter and temperature sensor.
Answer: This information has been supplied.
Comment 4.- Since the experiment was performed in the 2 specific months (when talking about temperature and light) can the authors provide more details about temperature control (especially heating) of the device. And what about light source when light is limited.
Answer: Temperature control has been used for cooling only. Details are given in M&M section. Results shown in this work are from month of March. In southern latitudes in Spain (Huelva), March is a month of moderate maximum and minimum temperatures, approximately 21°C (maximum) and 9°C (minimum) in average, respectively. Temperature increases rapidly after the sunrise. A new paragraph has been included in page 8: “heating was not implemented during the night and the outdoor cultures were produced under natural light-dark cycles. The productivity should be expected to increase in cultures subjected to 24h-illumination and temperature control, thus implementing artificial light and heating overnight. Nevertheless, these experiments should be carried out once the impact on process economy has been assessed”.
Comment 5.- Figure 2 – I would suggest putting same scale (from 0-1.8) on figures A and C for better visualisation and to correct x-axe on figure C so zero of both axes intersect.
Answer: Figure 2 has been modified accordingly.
Comment 6.- Conclusion is missing
Answer: Conclusions are now included in the text after the results and Discussion section.
Comment 7.- There are no newer references ( 5 are from 2016 y and the rest are older) so please provide newer.
Answer: Several recent references have been provided in the reference list.
Comment 8.- Please check English language.
Answer: With the exception of the new lines added to the text in the review process, the submitted version of the manuscript had been previously reviewed by a professional editing service (Sci-Edit).
Comment 9.- There are many small errors in the manuscript i.e.:
- row 122 …. dot at the end of the sentence is missing
Answer: It has been corrected.
- row 145 …. there is one excess bracket in the sentence
Answer: It has been corrected.
- rows 182 – 186 … some units in the paper are written with the · between them and most of them without so authors must decide what type they will use
Answer: It has been corrected.
- row 257…. C. should be in italic etc
Answer: It has been corrected.
Round 2
Reviewer 2 Report
Based on provided answers I feel that quality of the paper improved and that the manuscript is now suitable for publication
