Current Advances in Carotenoid Production by Rhodotorula sp.
, Rodolfo Ramos-González 3, Evelyn Faife-Pérez 4, Lourdes Georgina Michelena-Álvarez 4, José Luis Martínez-Hernández 1 and Anna Iliná 1,*Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsThe manuscript refers to the latest advances in terms of biotechnological carotenoid production by Rhodotorula sp yeast. It is well structured by first presenting the carotenogesis in Rhodotorula yeast and then, the main parameters that influence carotenoid production. Throughout the manuscript, it were observaed the latest references within this topic.
Comments for author File: 
Comments.pdf
It is well written, with very minor aspects to correct.
Author Response
Responses to Reviewer 1
I would like to thank you for your review and helpful observations of our manuscript Fermentation-2920954, entitled "Current advances in carotenoid production by Rhodotorula sp." Your constructive suggestions for improving our paper have been duly accepted and listed below. All text changes were marked in color.
Reviewers' comments:
Reviewer #1: Comments and Suggestions for Authors
The manuscript refers to the latest advances in terms of biotechnological carotenoid production by Rhodotorula sp yeast. It is well structured by first presenting the carotenogesis in Rhodotorula yeast and then, the main parameters that influence carotenoid production. Throughout the manuscript, it was observed the latest references within this topic.
It is well written, with very minor aspects to correct.
Comments
- Please add the name of the cycle
Response: tricarboxylic acid (TCA)
- remove the space
Response: This was modified as suggested, C/N=80
- Add capital letter - 'Potato' and uniform through the column
Response: The error was corrected: Potato wastewater /glycerol
- Capital letter
Response: This has been done: Hydrolyzed sugarcane molasses at 40 g/L
- Why is it in Italic letters? please Uniform through the column
Response: This has been changed now: Onion peel powder and mung bean husks
- Italics; please uniform
Response: This was changed as suggested: Sisal bagasse hydrolyzate Total carotenoids (223.5 μg/g) Batch, 28 °C, pH 6.0, 150 rpm, 120 h [44]
- Add space. Please uniform through the manuscript It should be written: ' 20-30 ºC Response: This was changed as suggested,
The best temperature for obtaining the highest biomass and carotenoid production is around 20-30 °C. Above 30°C, the synthesis of carotenoids decreases, with 25 °C being the ideal temperature for the maximum synthesis of these pigments [39, 52]. Kot et al. [53] reported a high increment in carotene biosynthesis by R. gracilis (360.4 µg/g), R. glutinis (280.3 µg/g), and R. mucilaginosa (150 µg/g) at 20 °C. Sharma & Ghoshal [9] optimized the production of carotenoids by R. mucilaginosa using agro-industrial wastes at 25.8 °C as the ideal temperature. Da Silva et al. [39] obtained a maximum carotenoid content (1.13 g/L) with R. mucilaginosa at 22 °C, while the lowest carotenoid (0.34 g/L) was obtained at 34 °C. Allahkarami et al. [44] indicated the increment in temperature (up to 28 °C) is accompanied by rising carotenoid formation (2.43 mg/L) by R. mucilaginosa while it was reduced above 35 °C (0.60 mg/L). Maia et al. [47] demonstrated that R. glutinis produced carotenoids at 35 °C.
- I suggest you to review this setence, it is not clear. You say that at low pH, the yeasts are forced to synthetize carotenoids, considering its high concentration, it that it? I only think the way is written is confusing. You could explain in 1 or 2 sentences, why low pH induce carotenoid production?
Response: Thank you very much for your comment. The changes were made.
The pH of the medium was changed according to the yeast growth and carotenoid production. During the synthesis of carotenoids in fermentation processes, there is a natural change in pH in the culture medium depending on the growth phase of the yeast. In general, pH decreases by the first 72 h; then, pH values increase because of the intensive carotenogenesis phase [51]. In general, Rhodotorula sp. prefers more acidic pH values; however, at shallow pH values, yeast growth is inhibited, resulting in the reduction of carotenoid production. Alkaline pH was suggested to act as a stressor and alter metabolic rates and nutrient uptake, inducing glucose cellular metabolism and enhancing polysaccharide synthesis instead of carotenoids. Xie et al. [26] proposed that a strategy to increase carotenoids is using two stages of fermentation with changes in pH: a first stage with a pH that favors growth and a second stage with a decrease in pH that stimulates the biosynthesis of carotenoids.
The highest production of carotenoids in Rhodotorulas sp has been reported in the pH ranges 5.0-6.0. At low pH, the increase of catalase and superoxide dismutase enzyme activity has been demonstrated in response to the induction of oxidative stress [7]. Da Silva et al. [39] reported that yeast growth was slow at a pH of approximately 5.0 and that carotene production was favored at this pH.
- Please the name with the respective acronym
Response. This was modified as suggested.
Lin et al. [75] used the genetic transformation of Agrobacterium tumefaciens to randomly insert the transfer DNA (T-DNA) containing hygromycin-resistant cassettes into the genome of R. toruloides NP11.
- dot
Response. It was corrected, Submerged fermentation strategies.
Author Response File: Author Response.pdf
Reviewer 2 Report
Comments and Suggestions for Authors- The abstract should be improved. The abstract should be more informative and descriptive.
- The aim of the study should be rephrased, as well as, the statement of the novelty should be provided. I wonder how this review article differs from others that have been written over the last few years. Please show what is new in this article.
- I propose to add the structural formulas of the described carotenoids.
- In my opinion, the section: „4. Genetic engineering to improve carotenoid production.” should be improved. A dozen papers were omitted while describing the possibilities of using genetic engineering in carotenoid production improvement.
- The reference list should be improved according to the instructions for authors (https://www.mdpi.com/journal/fermentation/instructions).
Author Response
Responses to Reviewer 2
I would like to thank you for your review and helpful observations of our manuscript Fermentation-2920954, entitled "Current advances in carotenoid production by Rhodotorula sp." Your constructive suggestions for improving our paper have been duly accepted and listed below. All text changes were marked in color.
Reviewer #2: Comments and Suggestions for Authors
- The abstract should be improved. The abstract should be more informative and descriptive.
Answer: Thank you very much for your comments, the abstract was modified according to your suggestions.
Microbial carotenoids are pigments of lipophilic nature; they are considered promising substitutes for chemically synthesized carotenoids in the food industry. The benefits for human health have been demonstrated due to its antioxidant capacity. Yeasts of the genus Rhodotorula have genotypic characteristics that allow them to accumulate high concentrations of carotenes under certain stress conditions. The present review includes recent information covering different aspects of carotenoid production in the Rhodotorula sp fermentation. The review focuses on fermentation carotenoid production strategies, describing various economic raw materials as a source of carbon and nitrogen, the capacity for tolerance to heavy metals, and the effect of light, pH, and salts on the accumulation of carotenoids. Genetic modification strategies to obtain strains with increased carotenoid production are described. Furthermore, using magnetic nanoparticles in the fermentation system, which could be a stress factor for increasing pigment production, is considered for the first time. Rhodotorula is a potential source of high-value carotenoids with applications in the cosmetics, pharmaceutical, and food industries.
- The aim of the study should be rephrased, as well as, the statement of the novelty should be provided. I wonder how this review article differs from others that have been written over the last few years. Please show what is new in this article.
Answer: The study's objective was reformulated, and the statement of novelty was provided.
This review describes the metabolic mechanism of carotenoid biosynthesis by Rhodotorula sp., and the influence of environmental factors as strategies to increase carotenoid production. Studies of genetic modification strategies are presented. For the first time, using magnetic nanoparticles in fermentation has been addressed as a strategy for accumulating secondary metabolites.
- - I propose to add the structural formulas of the described carotenoids.
Answer: Thank you for your comment. We have been presented the figure in accordance with your recommendation.
Figure 1. Structural formulas of several different kinds of carotenoids in the yeast genus Rhodotorula
4 - In my opinion, the section: "4. Genetic engineering to improve carotenoid production." should be improved. A dozen papers were omitted while describing the possibilities of using genetic engineering in carotenoid production improvement.
Answer: Thank you very much for your suggestion. We have enriched the section on genetic engineering. Figure 3 was changed to Table 2, a summary of several studies. At least 12 references to recent studies were included.
5 - The reference list should be improved according to the instructions for authors (https://www.mdpi.com/journal/fermentation/instructions).
Answer: Thank you very much for your suggestions. All the reference list was adjusted to MPDI Format.
Author Response File: Author Response.pdf
Round 2
Reviewer 2 Report
Comments and Suggestions for AuthorsThe manuscript has been carefully revised and all my comments were addressed.
