From Herbal Teabag to Infusion—Impact of Brewing on Polyphenols and Antioxidant Capacity

Round 1

Reviewer 1 Report

This is a very interesting and practical paper on how brewing of teas will influence the availability of specific phytochemicals and the associate antioxidant activity provided by the mixture of available phytochemicals. 

The experiments are well designed and carried out.  The work is well done with replicates and the paper is very well written and easy to read.

My major and only criticism lies with the Figures 2, 3.  For figure 2, it is hard to differentiate the difference between FPC and TFC on the graph.  I also do not understand the meaning of the shared letters as the hybrid ones contain letters that are not singly identified (e.g. n, b etc.).  What does np mean then?  The same is true for Figure 3 where at 11 minutes we have a c, bc, c denoted differences, but no single b..  Hence, I do not understand the significance of this MRT.    Please double check the statistical analyses. 

Was Figure 11 chromatogram data supported by individual standards as well as a mixture of these particular standards.  The RT's for some of them are very close and the significant figures on the elution times is questionable.  The principle peaks are not the flavonoids identified, so do the authors know what they are, since they are major components of the chromatogram

Author Response

This is a very interesting and practical paper on how brewing of teas will influence the availability of specific phytochemicals and the associate antioxidant activity provided by the mixture of available phytochemicals. 

The experiments are well designed and carried out.  The work is well done with replicates and the paper is very well written and easy to read.

The Authors would like to thank the Reviewer for the constructive comments.

My major and only criticism lies with the Figures 2, 3.  For figure 2, it is hard to differentiate the difference between FPC and TFC on the graph.  I also do not understand the meaning of the shared letters as the hybrid ones contain letters that are not singly identified (e.g. n, b etc.).  What does np mean then?  The same is true for Figure 3 where at 11 minutes we have a c, bc, c denoted differences, but no single b..  Hence, I do not understand the significance of this MRT.    Please double check the statistical analyses. 

We have revised Figure 2 to better show the difference between TPC and TFC. Regarding to the letters, which statistically differentiate between different conditions, we have checked and revised the letters in Figure 2. In addition, we have also revised the legends of all Figures to make them clearer.

Was Figure 11 chromatogram data supported by individual standards as well as a mixture of these particular standards.  The RT's for some of them are very close and the significant figures on the elution times is questionable.  The principle peaks are not the flavonoids identified, so do the authors know what they are, since they are major components of the chromatogram

Figure 11 chromatogram data were supported by individual standards as well as a mixture of these standards. We not only ran individual standards as well as a mixture of these standards to compare the retention time with the peaks of the herbal tea extract but also mixed each individual standard to the herbal tea extract to observe that specific peak area high with corresponding retention time in order to double confirm the retention time of each individual compounds. Therefore, the compounds and their RT showed in the manuscript are accurate. Some major peaks have not been identified and we do not know what they are. We will isolate and identify them in future studies.

Reviewer 2 Report

The authors investigated the impact of brewing on antioxidant capacity and phenolic content in herbal teabag infusions. The authors presented data comparison using essential 3 types of tea infusion techniques and a laboratory extraction. The infusions were analyzed for total phenolic and flavonoid content. Additionally, three antioxidant activity assays were performed on the tea infusions. Furthermore, a qualitative evaluation of the phytochemical diversity was presented using an HPLC-PDA analysis, including comparisons of 7 analytical reference standards of know phytochemicals. Overall the manuscript is worthy of publication after some minor revisions and clarifications.

Minor comments:

Line 132: Include the purity and vendor info of gallic acid standard used.

Line 140: Include the purity and vendor info of the catechin used.

Line 163: Dimensions of HPLC column should be 3 x 250 mm

Line 140: Include the purity and vendor info of the Trolox used.

Line 170: Include information on the analytical standards used as an RT reference.

Line 368: the word “identified” may not be entirely accurate as some of these compounds may not have standards available. It is likely more appropriate to convey to the reader that there are over 20 major chromatographic peaks, of which 7 were identified using reference standards.

Line 371-373: I would reduce the significant digits on the retention times reported.

Line 390: The statement that there are 30 individual compounds identified is not accurate. No table listing 30 unique compounds that have been appropriately identified is presented.

Figures: Generally, be consistent with the formatting throughout the manuscript, and keep axes labels and font formatting the same.

Author Response

The authors investigated the impact of brewing on antioxidant capacity and phenolic content in herbal teabag infusions. The authors presented data comparison using essential 3 types of tea infusion techniques and a laboratory extraction. The infusions were analyzed for total phenolic and flavonoid content. Additionally, three antioxidant activity assays were performed on the tea infusions. Furthermore, a qualitative evaluation of the phytochemical diversity was presented using an HPLC-PDA analysis, including comparisons of 7 analytical reference standards of know phytochemicals. Overall the manuscript is worthy of publication after some minor revisions and clarifications.

The Authors would like to thank the Reviewer for the favourable comments.

Minor comments:

Line 132: Include the purity and vendor info of gallic acid standard used.

The purity and vendor info of gallic acid standard have been added.

Line 140: Include the purity and vendor info of the catechin used.

Purity and vendor info of the catechin have been added.

Line 163: Dimensions of HPLC column should be 3 x 250 mm

Dimensions of HPLC column have been corrected.

Line 140: Include the purity and vendor info of the Trolox used.

Purity and vendor info of the Trolox have been included.

Line 170: Include information on the analytical standards used as an RT reference.

Information on the analytical standards has been included.

Line 368: the word “identified” may not be entirely accurate as some of these compounds may not have standards available. It is likely more appropriate to convey to the reader that there are over 20 major chromatographic peaks, of which 7 were identified using reference standards.

We have revised the manuscript as suggested by the Reviewer.

Line 371-373: I would reduce the significant digits on the retention times reported.

We have reduced the significant digits as suggested by the Reviewer.

Line 390: The statement that there are 30 individual compounds identified is not accurate. No table listing 30 unique compounds that have been appropriately identified is presented.

We have revised the manuscript as suggested by the Reviewer. It now reads “There are 20 major chromatogram peaks, of which 7 were identified using reference standards, revealing that this herbal tea can link with health benefits.”

Figures: Generally, be consistent with the formatting throughout the manuscript, and keep axes labels and font formatting the same.

We have revised the Figures with the consistent format as suggested by the Reviewer.