Development of Functional Fermented Dairy Products Containing Taiwan Djulis (Chenopodium formosanum Koidz.) in Regulating Glucose Utilization

Round 1

Reviewer 1 Report

Fermentation Taiwan Djulis

Authors presented their study of adding Taiwan Djulis during yogurt fermentation process and studies the effects of fermented product on glucose utilization. Comments are listed as below:

1.       Line 24, the objective of this study was to develop fuctional Taiwan Djulis fermented dairy products. However, the author only developed one product, and whether it can be claimed as functional dairy product, is there any guideline on functional product? It is not clearly described in this study.

2.       Line 28-line 32, are these results are for the fermented Djulis dairy product? How much higher than control sample, fermented milk without Djulis?

3.       Line 45, Total phenols, is it more appropriate to call it total polyphenol compounds? Please keep the same name throughout this manuscript.

4.       Line 49, “gained importance”, change another word?

5.       Line 71, what is shelled djulis? Do you mean deshelled or with shell? In the fermentation, did you add whole grain, or whole grain powder? Or cooked grain? Is it raw? Is it edible as raw?

6.       Line 142, only fermentation of Djulis is mentioned, how about control? Fermentation without djulis? What is the fermentation medium? Did you do plate count to check the growth curve throughout the fermentation? How do you determine the time point to stop the fermentation? Did you add other nutrients during fermentation?

7.       Line 251, move reference number to be after “antioxidant capacity”.

8.       Line 254, djulis fermented dairy products, what are the products? It is better to say” fermented djulis dairy product or products”?

9.       Line 274-275, move reference not [28,29] to be after “respectively”.

0.   Line 290. This result is obvious, the more you add and the higher value of antioxidative active you will get. The problem is whether different amount of djulis added will affect the growth of the strains, the taste of the product, the appearance of the product and so on.

1.   Line 306, same comments as above, this is obvious.

1.   Line 325, post addition of djulis? Do you mean you add it after fermentation? This is not stated clearly at method section.

1.   Line 329 to line 333, line 328 said no significant difference on boned polyphenol content, then at line 329 you said during fermentation bacteria will have enzyme to hydrolyze to release phenolic compound, then boned phenolic compounds will be less. That is not reflected in your result.

1.   Line 366, only the result was put here without discussion. Is this amount of GABA sufficient to create health effect at certain intake amount?

1.   Line 425 what do you mean the acid was 5.3 X 108 …?

1.   Line 426, storage will let the cell count drop, shouldn’t you measure the cell count daily? After 14 days how much cell do you have in the product?

1.   Line 427-428, you store it at 4 degree, do you expect them to grow during storage? Is not it because of pH but the temperature?

1.   Figure 1, what do mean on the legends? Could you describe them in the figure title?

1.   Line 444, you did fermented milk but not yogurt? Why do you compare it with yogurt?

2.   Line 484-486, no significance was observed, why you said glucose utilization can be increased to 1.6 times? What is your control? Did you do control with ethanol?

2.   Figure S2, put figures in the same order as it was presented in the title, put (a), (b) (c) on the graph.

2.   Figure S3, no significant difference was observed in (a), and survival of cells was lower than control in (b). that is not a good result, does it mean there is no effect at all?

Author Response

Authors presented their study of adding Taiwan Djulis during yogurt fermentation process and studies the effects of fermented product on glucose utilization. Comments are listed as below:

Thanks to the reviewer's constructive suggestion to improve the quality of this manuscript, the author would give the highest respect.

1. Line 24, the objective of this study was to develop fuctional Taiwan Djulis fermented dairy products. However, the author only developed one product, and whether it can be claimed as functional dairy product, is there any guideline on functional product? It is not clearly described in this study.

Thanks for your suggestion. We add ‘‘ and use FL83b cell to evaluate the glucose utilization ability ’’.in line 28, and delete ‘‘ functional ’’ in line 27. We used L. plantarum, L. bulgaricus and S. thermophilus to develop the fermented products. That strain was a traditional microbial composition in Set yogurt or Drinking yogurt and could change the prototype product by adjusting fermented time. And we further test the potential bioactivity on regulating glucose utilization, therefore we claimed as functional dairy product.

2. Line 28-line 32, are these results are for the fermented Djulis dairy product? How much higher than control sample, fermented milk without Djulis?

In line 28-line 32, all the results are belonging fermented Djulis dairy product. In order to described clearly on abstract, we add ‘‘ Compare to fermented milk without Djulis ’’ in line 32, and present the growth rate in line 34, line 35, and line 36.

3. Line 45, Total phenols, is it more appropriate to call it total polyphenol compounds? Please keep the same name throughout this manuscript.

We revise this mistake in line 51. Thanks for your recommendation.

4. Line 49, “gained importance”, change another word?

We replace “gained importance” to ‘‘ become important ’’ in line 55.

5. Line 71, what is shelled djulis? Do you mean deshelled or with shell? In the fermentation, did you add whole grain, or whole grain powder? Or cooked grain? Is it raw? Is it edible as raw?

On this study, the fermented substrates are deshelled form of djulis. It can’t edible as raw, due to raw djulis have anti-nutrient factor (phytic acid and saponin), there djulis need to cook or ferment to reduce it. We revise ‘‘shelled form’’ to ‘‘deshelled’’ in line 76 and line 77. Thanks for your modification opinions.

6. Line 142, only fermentation of Djulis is mentioned, how about control? Fermentation without djulis? What is the fermentation medium? Did you do plate count to check the growth curve throughout the fermentation? How do you determine the time point to stop the fermentation? Did you add other nutrients during fermentation?

We are sorry for the imprecise expression; the substrate of control group is fresh milk without adding djulis and don’t add any other nutrients. We actually used plate counting to determine the growth curve but data didn’t show in this article, therefore we revise the description of this section in line 153-165.

7. Line 251, move reference number to be after “antioxidant capacity”.

Thanks for your suggestion. We revise it in line 274

8. Line 254, djulis fermented dairy products, what are the products? It is better to say” fermented djulis dairy product or products”?

We revise ‘‘djulis fermented dairy products ’’ to ‘‘fermented djulis dairy product’’ in line 286.

9. Line 274-275, move reference not [28,29] to be after “respectively”.

Thanks for your suggestion. We revise it in line 306.

10. Line 290. This result is obvious, the more you add and the higher value of antioxidative active you will get. The problem is whether different amount of djulis added will affect the growth of the strains, the taste of the product, the appearance of the product and so on.

We believe that the concentration of djulis will affect many aspects of the final fermented product, however articles are limited in length. We choose to percent the changes of bioactive property (such as antioxidant and nutrient composition) throughout of fermentation, meanwhile focus on the functional ability of our fermented products. Thanks for your suggestion, we will determine those experience in future work.

11. Line 306, same comments as above, this is obvious.

We reply this recommend above. Thank you.

12. Line 325, post addition of djulis? Do you mean you add it after fermentation? This is not stated clearly at method section.

We delete ‘‘ post-addition ’’ in line 361. Thank you.

13. Line 329 to line 333, line 328 said no significant difference on boned polyphenol content, then at line 329 you said during fermentation bacteria will have enzyme to hydrolyze to release phenolic compound, then boned phenolic compounds will be less. That is not reflected in your result.

We revise ‘‘ was not significantly ’’ to ‘‘ and the fraction bound to phenol shown significantly decreased 13.65% (92.78±0.88 mg GAE g−1) ’’ in line 364, 365. Thanks for your suggestion.

14. Line 366, only the result was put here without discussion. Is this amount of GABA sufficient to create health effect at certain intake amount?

Yu et al. developed GABA-rich fermented milk, and test the once-daily oral doses of GABA-fermented milk (8.83~ 33.33 mg/kg.bw) on mice, shown the beneficial effect on insomnia involving regulation of gut microbiota. We add this description in line 405 and 407 and renew it on the reference [41].

15. Line 425 what do you mean the acid was 5.3 X 108 …?

We are sorry for the imprecise expression. We revise line 469 – 473, show the change of the pH value throughout the storage period.

16. Line 426, storage will let the cell count drop, shouldn’t you measure the cell count daily? After 14 days how much cell do you have in the product?

We follow the research on Journal of Dairy Science (Salvador and Fiszman., 2004). In long storage test, the research team determined the cell count at day 7, 14 and 21. The amount of cell were 5.3 X 10⁸ on day 14. https://doi.org/10.3168/jds.S0022-0302(04)73544-4

17. Line 427-428, you store it at 4 degree, do you expect them to grow during storage? Is not it because of pH but the temperature?

In long storage test, we simulate fermented dairy products stored on the shelf at 4 degree. The titratable acidity increasing and pH value decreasing reaveled that the content of organic acid in fermented products raises up to reduce the strain survival. Meamwhile low temperature will also affect the viable count. We revise line 457-458 to express this opinion.

18. Figure 1, what do mean on the legends? Could you describe them in the figure title?

We add the abbreviation of the four fermented products. In this section, our research team recruit 52 untrained consumer to join this sensory evaluation. We revise this section from line 502 to 506.

19. Line 444, you did fermented milk but not yogurt? Why do you compare it with yogurt?

As suggestion 1 mentioned, we develop fermented dairy products, our fermented products shown more softer texture than traditional yogurt and only partially curdled, however as a comparison product for sensory evaluation, commercial yogurt is closer to our fermented product.

20. Line 484-486, no significance was observed, why you said glucose utilization can be increased to 1.6 times? What is your control? Did you do control with ethanol?

   The control group were cell without treatment. We revise the description in line 526 – 528.

21. Figure S2, put figures in the same order as it was presented in the title, put (a), (b) (c) on the graph.

       Thanks for your suggestion. We add the (a), (b) (c) on the graph.

22. Figure S3, no significant difference was observed in (a), and survival of cells was lower than control in (b). that is not a good result, does it mean there is no effect at all?

Figure S3 were evaluate the toxicity of the ethanol extracts in different dose. And all the extraction shown no effect on cell survival (survival > 80%).

 

 

Reviewer 2 Report

The strength of this article is represented by the concept of research and development in the field of nutraceuticals using relatively simple technologies such as fermentation, but optimized in order to obtain increasingly specific and healthy products for the population. My compliments to the research team. A small nite related to the continuation of the work, is to approach a clinical study on patients with reduced glucose tolerance in order to evaluate the functionality of the product studied. I hope to see the product on the market soon.

Author Response

The strength of this article is represented by the concept of research and development in the field of nutraceuticals using relatively simple technologies such as fermentation, but optimized in order to obtain increasingly specific and healthy products for the population. My compliments to the research team. A small nite related to the continuation of the work, is to approach a clinical study on patients with reduced glucose tolerance in order to evaluate the functionality of the product studied. I hope to see the product on the market soon.

 

Thanks to the reviewer's constructive suggestion to improve the quality of this manuscript, the author would give the highest respect.

 

Reviewer 3 Report

Dear authors

this manuscript shows a novel idea, however unfortunately it is not well designed and presented. 

- First, the English language of the manuscript is poor 

-Please rewrite the title it could be way better

- the abstract should be rewritten. it should be as clear and informative as possible

-Where is the method for sensory evaluation???!! you have results for sensory evaluation while there is no methodology in this manuscript!!!

- The manuscript is not well  designed at all, especially in the materials and methods section. 

-you have mentioned fermented dairy products several times. What kind of products? even in table 1 you have compared the composition and biological activity of fermented dairy products and Taiwan djulis fermented dairy products. you should first explain what kind of dairy product are you talking about.

-Why the figure 1 is presented before its related results???

- In figure 1, the SD of measured parameters are so high. this means there has been a high variation in your data. why is that?

Author Response

Dear authors

this manuscript shows a novel idea, however unfortunately it is not well designed and presented. 

Thanks to the reviewer's constructive suggestion to improve the quality of this manuscript, the author would give the highest respect.

- First, the English language of the manuscript is poor 

Thanks for your suggestion. We revise the English language of the manuscript.

-Please rewrite the title it could be way better

Thanks for your suggestion. We revise it in line 2-4.

- the abstract should be rewritten. it should be as clear and informative as possible

We revise it in line 27-40. Thank you.

-Where is the method for sensory evaluation???!! you have results for sensory evaluation while there is no methodology in this manuscript!!!

We add the methodology in line 236 to 248. Thanks for your recommendation.

- The manuscript is not well designed at all, especially in the materials and methods section.

We revise the descriptions of ‘‘2.5 Fermentation and fermentation broth freeze drying’’ and add the methodology of ‘‘Sensory evaluation’’. Thanks you.

-you have mentioned fermented dairy products several times. What kind of products? even in table 1 you have compared the composition and biological activity of fermented dairy products and Taiwan djulis fermented dairy products. you should first explain what kind of dairy product are you talking about.

We develop fermented dairy products, our fermented products shown more softer texture than traditional yogurt and only partially curdled. The ‘‘fermented dairy products’’ means fermented dairy without Taiwan djulis. We add the description in Table1 legends in line 346.

-Why the figure 1 is presented before its related results???

We revise this mistake in line 501. Thanks for your recommendation.

- In figure 1, the SD of measured parameters are so high. this means there has been a high variation in your data. why is that?

In this part of the experiment, we recruited 53 untrained general consumers, which may make the score description slightly imprecise. However, it also reflects that consumers are likely to have little difference in these fermented products, showing that our fermented products are acceptable to consumers and have a certain degree of competitiveness.

 

Round 2

Reviewer 3 Report

Thanks for preparing this manuscript as it is worth to publish after adding a conclusion phrase at the end of the abstract.

Author Response

Thanks for your suggestion, we add ‘‘This functional fermented dairy product can be developed into high added value of local Agri-cultural products and enhance multiple applications.’’ at the end of the abstract in line 36-37.