Quantitative Assessment of Cold Injury in Tea Plants by Terahertz Spectroscopy Method

Round 1

Reviewer 1 Report

Authors propose a non-destructive method to assess the Cold injury of tea leaf based on the terahertz (THz) spectroscopy.

It is an interesting research but authors should fix some issues to have the paper accepted:

- The structure of the paper should be explained at the end of the introduction section.

- Authors should compare their method with existing systems. They should provide a commparison table at least.

- Authors should include their futture work at the end of the conclusion section.

Author Response

Dear reviewer,

Thank you for your letter and the reviewers’ comments concerning our manuscript entitled: “Quantitative Assessment of Cold Injury for Tea by Terahertz Spectroscopy Method”. (ID: agronomy-2388089). Thank you for giving our opportunity to revise our manuscript. Those comments are all valuable and very helpful for improving our manuscript. We have studied the comments carefully and have made correction which we hope will meet your request and for approval.

The reviewer’s comments are marked in yellow and our responses are marked in green.

Best,

Yongzong Lu （[email protected]） and Yongguang Hu ([email protected])

Q1: Authors propose a non-destructive method to assess the Cold injury of tea leaf based on the terahertz (THz) spectroscopy. It is an interesting research but authors should fix some issues to have the paper accepted.

R: Thanks for your interest on this research. I have made the necessary changes accordingly. Thank you again.

Q2：The structure of the paper should be explained at the end of the introduction section.
R: Thanks for your kind comments. I have added the structure of the paper. The relevant corrections have been made in revised version in the manuscript as following:

“The remainder of this paper is organized as follows: detailed materials and methods used in this research were presented in section 2; related results and discussions were provided in section 3; the conclusions of the overall paper and future research points were proposed in section 4.”

The relevant corrections have been made in revised version in the manuscript.

Q3: Authors should compare their method with existing systems. They should provide a comparison table at least.
R: Thanks for your kind comments. Various techniques have been developed to monitor and determine the degree of CI in plants. The destructive technique some of which involves clamping can damage the leaf depending on the clamping force and the clamping duration which may render the data acquired defective. Moreover, they are time consuming, and not suitable for on-line detection. Existing systems or techniques have been employed with the specific objective of monitoring the changes in the macro and micro structure of the plant tissue to assess the degree of cold damage and cold tolerance. In this research, we were focused on providing a new quick non-destructive method is provided to assess the CI of tea leaf based on the terahertz (THz) spectroscopy. In our future research, we will focus on the evaluating different methods.

The relevant corrections have been made in revised version in the manuscript.

Q4: Authors should include their future work at the end of the conclusion section.

R: Thanks for your kind comments on improving the quality of my manuscript to be better accepted. We accept you kind comments and add the future work at the end of conclusion section.

“In the future research, we will evaluate the THz spectroscopy and Hyperspectral imaging techniques on detecting freezing injury for other plants that are grown in freeze/frost prone area. Further research will also focus on the improvement in assessing the cold damage should include at least -15 °C and if possible up to -25 °C since in the plant environment, the plant canopy temperature can sometime decrease to as low as -30 °C. This will ensure that data is made available for all seasons and all regions and seasonal fluctuations in the ambient temperature. ”

The relevant corrections have been made in revised version in the manuscript.

Author Response File: Author Response.pdf

Reviewer 2 Report

Quantitative Assessment of Cold Injury for Tea by Terahertz 2 Spectroscopy Method.

The paper describes a new method for determining cold injury in tea leaves.

The authors explain that THz spectroscopy must be combined with 2DCOS-PLSR model with chemometrics to give the most accurate results of cold injury to tea leaves.

It is important to be able to accurately determine cold injury in tea leaves in order to search for ways in cold tolerance in tea leaves could be improved.

The description of the methods is clear. The biggest problem is that the figures and table is not close to where it is first mentioned, therefore making it difficult to follow. In line 257 is mention of Figure 2b, which couldn’t be found.

The language is good but could be improved.

The paper describes a novel way of assessing cold injury in leaves and could find application in other studies. Therefore, it is a paper which significantly contributes to the field.

The language in the introduction is good, but the quality of the language drops in the Materials and Methods. Then it improved again in the conclusion. A problem which could be fixed fairly easily with the help of an english editor.

Author Response

Dear reviewer,

Thank you for your letter and the reviewers’ comments concerning our manuscript entitled: “Quantitative Assessment of Cold Injury for Tea by Terahertz Spectroscopy Method”. (ID: agronomy-2388089). Thank you for giving our opportunity to revise our manuscript. Those comments are all valuable and very helpful for improving our manuscript. We have studied the comments carefully and have made correction which we hope will meet your request and for approval.

The reviewer’s comments are marked in yellow and our responses are marked in green.

Best,

Yongzong Lu （[email protected]） and Yongguang Hu ([email protected])

Q1: The paper describes a new method for determining cold injury in tea leaves. The authors explain that THz spectroscopy must be combined with 2DCOS-PLSR model with chemometrics to give the most accurate results of cold injury to tea leaves. It is important to be able to accurately determine cold injury in tea leaves in order to search for ways in cold tolerance in tea leaves could be improved.The description of the methods is clear. The biggest problem is that the figures and table is not close to where it is first mentioned, therefore making it difficult to follow. In line 257 is mention of Figure 2b, which couldn’t be found.

R: Thanks for your kind comments on improving the quality of my manuscript to be better accepted. We accept your kind comments. We are sorry to make several mistakes in the line 257 where mentioned the Figure 2b. We have deleted the figure 2b in the manuscript and we also checked all the figures and tables to make sure the quotations are all correct.

The relevant corrections have been made in revised version in the manuscript.

Q2: The language is good but could be improved.The paper describes a novel way of assessing cold injury in leaves and could find application in other studies. Therefore, it is a paper which significantly contributes to the field. The language in the introduction is good, but the quality of the language drops in the Materials and Methods. Then it improved again in the conclusion. A problem which could be fixed fairly easily with the help of an english editor.

R: Thanks for your kind comments. We have carefully checked the English language of the whole manuscript and proofread by two native speakers before resubmitted.

The relevant corrections have been made in revised version in the manuscript.

Author Response File: Author Response.pdf

Reviewer 3 Report

The manuscript describes the application of a non-destructive method based on the terahertz (THz) spectroscopy in assessing the cold injury of tea leaf, combined with 2DCOS-PLSR model toward providing a better benchmark for the input interval selection and improved accuracy of cold injury detection result.

The topic of applying THz and chemo metric methods for improved and more efficient data analysis is novel and of practical significance. The scope is adequate and the methods are appropriate.

However, the Introduction does not provide sufficient background of the THZ technique for uninitiated readers to follow. The reproducibility is relatively low in that some key operating parameters are missing, such as THZ spectral resolution, acquisition rate and dynamic range. Section 2.3 is routine practice of THz experiments and are generally understood. If the authors insist on presenting it with details, please move it to Supplementary information, or simply mention it at the end of section 2.2.

What the result of the “Quantitative Assessment of Cold Injury for Tea” is appears unclear though, please excuse this old reviewer’s failing eyes. Please present the result in a more straightforward fashion, for example which temperature affects the tea most, in which THz range shows the most characteristics changes compared to the control, by what %, etc.

Moderate English check is needed.

Thus a major revision is recommended before further consideration.

Some suggestions:

1. To remedy the lacking of background of the THz technique, the authors may refer to the following 2 publications where these have been introduced.

a. Song FV, Yang B, Di Tommaso D, Donnan RS, Chass GA, Yada RY, Farrar DH, Tian KV. Resolving nanoscopic structuring and interfacial THz dynamics in setting cements. Materials Advances. 2022;3(12):4982-90.

b. Tian KV, Yang B, Yue Y, Bowron DT, Mayers J, Donnan RS, Dobó-Nagy C, Nicholson JW, Fang DC, Greer AL, Chass GA. Atomic and vibrational origins of mechanical toughness in bioactive cement during setting. Nature communications. 2015 Nov 9;6(1):8631.

2. Is tea also used for food? Change drink to beverage.

3. Check the English, “…Long Jing with five years old…” change to a five-year-old Longjing; change “the rest of water on leaves were moped with blotting paper” to were blotted with blotting paper.

4. Fig. 1 caption does not introduce panel a-c.

5. Some equations do not have references, unless they are original.

6. Fig. 2, can you also plot them in 2 D, absorbance-frequency plot, offset the 5 plots for guiding the eye and simply mark the temperature next to each curve, in order to present which temperature affects the tea most, in which THz range shows the most characteristics changes compared to the control, and if you could quantify this change? Just a suggestion, maybe you have better ways of presenting the results of the Quantitative Assessment of Cold Injury for Tea.

7. Table 1 does not have RMSE_2D and if the authors want to say “a significant increase in R_2D, a statistical analysis is needed here.

moderate modification

Author Response

Dear reviewer,

Thank you for your letter and the reviewers’ comments concerning our manuscript entitled: “Quantitative Assessment of Cold Injury for Tea by Terahertz Spectroscopy Method”. (ID: agronomy-2388089). Thank you for giving our opportunity to revise our manuscript. Those comments are all valuable and very helpful for improving our manuscript. We have studied the comments carefully and have made correction which we hope will meet your request and for approval.

The reviewer’s comments are marked in yellow and our responses are marked in green.

Best,

Yongzong Lu （[email protected]） and Yongguang Hu ([email protected])

Q1: The manuscript describes the application of a non-destructive method based on the terahertz (THz) spectroscopy in assessing the cold injury of tea leaf, combined with 2DCOS-PLSR model toward providing a better benchmark for the input interval selection and improved accuracy of cold injury detection result. The topic of applying THz and chemo metric methods for improved and more efficient data analysis is novel and of practical significance. The scope is adequate and the methods are appropriate. However, the Introduction does not provide sufficient background of the THZ technique for uninitiated readers to follow. The reproducibility is relatively low in that some key operating parameters are missing, such as THZ spectral resolution, acquisition rate and dynamic range. Section 2.3 is routine practice of THz experiments and are generally understood. If the authors insist on presenting it with details, please move it to Supplementary information, or simply mention it at the end of section 2.2.

R: Thanks for your kind comments on improving the quality of my manuscript to be better accepted. We have added the detailed key operating parameters of THZ spectral resolution, acquisition rate and dynamic rang in the end of section 2.2 which was as shown in table 1 and figure 1. We have also added the two recommended publications to remedy the lacking of background of the THz technique.

Table 1  Terahertz spectrum settings for the measurements

Figure 1. Plant under freezing in the MIR253 (a), Spectra measurement with THz-TDS (b) and structure of the THz-TDS system (c). The frequency resolution of TS7400 was set to 1.9GHz, the dynamic range was set to ＞ 50dB, acquisition rate was set to 16 ms/scan.

[39] Song FV, Yang B, Di Tommaso D, Donnan RS, Chass GA, Yada RY, Farrar DH, Tian KV. Resolving nanoscopic structuring and interfacial THz dynamics in setting cements. Materials Advances. 2022, 3(12), 4982-90.

[40] Tian KV, Yang B, Yue Y, Bowron DT, Mayers J, Donnan RS, Dobó-Nagy C, Nicholson JW, Fang DC, Greer AL, Chass GA. Atomic and vibrational origins of mechanical toughness in bioactive cement during setting. Nature communications. 2015, 6(1), 8631.

Q2: What the result of the “Quantitative Assessment of Cold Injury for Tea” is appears unclear though, please excuse this old reviewer’s failing eyes. Please present the result in a more straightforward fashion, for example which temperature affects the tea most, in which THz range shows the most characteristics changes compared to the control, by what %, etc.

R: Thanks for your kind comments on improving the quality of my manuscript to be better accepted. We are sorry to make you confused on the results and conclusion. As shown in the figure 2, the trend has shown that seven chief peaks and three subordinate peaks exist in the band. The chief peaks could be found in 0.21 THz, 0.46 THz, 0.62 THz, 0.78 THz, 1.0 THz, 1.32 THz and 1.54 THz and the subordinate peaks could be found in 0.32 THz, 0.89 THz and 1.2 THz. Average THz absorbance of tea plant samples with different low temperature stress generally increased significantly with the decrease in temperature. The trend for the temperature of 0°C is far lower than the trend for the other temperatures and could be discriminated easily. This is because serious injury to tea plants takes place at temperatures below 0°C. The differences in the absorption peaks for the other temperature levels could not be discriminated easily due to closeness and overlap. In this study, a new method of assessing the CI of a tea cultivar (Long Jing) by measuring the absorption spectra obtained from a non-distractive THz spectroscopy has been established. The study demonstrates the effectiveness of THz technique in the characterization of the changes associated with cold injury of tea leaves. Though studies have shown that substantial scattering of injured cells in the leaf results in overlap signals which restrict accurate assessment of damage cell absorption in the THz region, the com-bination of the THz spectroscopy technique and 2DCOS-PLSR model with chemometrics has provided feasible and effective quantitative assessment of cold injury in tea leaves.

Q3. To remedy the lacking of background of the THz technique, the authors may refer to the following 2 publications where these have been introduced.

R: Thanks for your kind comments on improving the quality of my manuscript to be better accepted. We have added the two recommended publications to remedy the lacking of background of the THz technique.

[39] Song FV, Yang B, Di Tommaso D, Donnan RS, Chass GA, Yada RY, Farrar DH, Tian KV. Resolving nanoscopic structuring and interfacial THz dynamics in setting cements. Materials Advances. 2022, 3(12), 4982-90.

[40] Tian KV, Yang B, Yue Y, Bowron DT, Mayers J, Donnan RS, Dobó-Nagy C, Nicholson JW, Fang DC, Greer AL, Chass GA. Atomic and vibrational origins of mechanical toughness in bioactive cement during setting. Nature communications. 2015, 6(1), 8631.

Q4: Is tea also used for food? Change drink to beverage.

R: Thanks for your kind comments. We have carefully checked the English language of the whole manuscript and proofread by two native speakers before resubmitted. We changed the drink to beverage.

The relevant corrections have been made in revised version in the manuscript.

Q5. Check the English, “…Long Jing with five years old…” change to a five-year-old Longjing; change “the rest of water on leaves were moped with blotting paper” to were blotted with blotting paper.

R: Thanks for your kind comments on improving the quality of my manuscript to be better accepted. We have carefully checked the English language and made relevant corrections.

The relevant corrections have been made in revised version in the manuscript.

Q6: Fig. 1 caption does not introduce panel a-c.

R: Thanks for your kind comments. We introduced the panel a-c as following.

Figure 1. Plant under freezing in the MIR253 (a), Spectra measurement with THz-TDS (b) and structure of the THz-TDS system (c). The frequency resolution of TS7400 was set to 1.9GHz, the dynamic range was set to ＞ 50dB, acquisition rate was set to 16 ms/scan.

The relevant corrections have been made in revised version in the manuscript.

Q7: Some equations do not have references, unless they are original.

R: Thanks for your kind comments. We have checked the whole manuscript and added all the references of the equations.

The relevant corrections have been made in revised version in the manuscript.

Q8: Fig. 2, can you also plot them in 2 D, absorbance-frequency plot, offset the 5 plots for guiding the eye and simply mark the temperature next to each curve, in order to present which temperature affects the tea most, in which THz range shows the most characteristics changes compared to the control, and if you could quantify this change? Just a suggestion, maybe you have better ways of presenting the results of the Quantitative Assessment of Cold Injury for Tea.

R: Thanks for your kind comments on improving the quality of my manuscript to be better accepted. We have revised the figure 2 to make sure it could clearly present the useful information. The trend has shown that seven chief peaks and three subordinate peaks exist in the band. The chief peaks could be found in 0.21 THz, 0.46 THz, 0.62 THz, 0.78 THz, 1.0 THz, 1.32 THz and 1.54 THz and the subordinate peaks could be found in 0.32 THz, 0.89 THz and 1.2 THz (Figure 2). Average THz absorbance of tea plant samples with different low temperature stress generally increased significantly with the decrease in temperature. The trend for the temperature of 0°C (control treatment) is far lower than the trend for the other temperatures and could be discriminated easily. This is because serious injury to tea plants takes place at temperatures below 0°C. The differences in the absorption peaks for the other temperature levels could not be discriminated easily due to closeness and overlap.

Figure 2. Absorbance spectra in the band of 0.1–1.6THz of the tea leaf response to different low temperature.

The relevant corrections have been made in revised version in the manuscript.

Q9: Table 1 does not have RMSE_2D and if the authors want to say “a significant increase in R_2D, a statistical analysis is needed here.

R: Thanks for your kind comments on improving the quality of my manuscript to be better accepted. We are sorry that we made the mistake on the Table 1. We accept your comments and made relevant revisions on the table 1 as following. Actually, in the Line 302 to Line 305, we described the information of quantitative assessment results “the quantitative assessment result obtained for the 2DCOS-PLSR, R_2D is 0.7873, 0.8305 and 0.9103, respectively. RMSE_2D is 0.6032, 0.5763 and 0.5221, respectively. For AI-PLSR model, R_AI is 0.7477, 0.7691 and 0.8974, respectively. RMSE_AI is 0.6038, 0.5962 and 0.5797, respectively.”

Table 1. Comparison of original PLSR and 2DCOS-PLSR performance

The relevant corrections have been made in revised version in the manuscript.

Author Response File: Author Response.pdf

Round 2

Reviewer 3 Report

The majority of the issues raised have been addressed.

Fig.2, is hard to see different plots. Offset each for clarity.

Fig. 4, 5 could be combined to show the improvement. Also please use the same x scale for Fig. 5b.

Author Response

Dear reviewer,

Thank you for your letter and the reviewers’ comments concerning our manuscript entitled: “Quantitative Assessment of Cold Injury for Tea by Terahertz Spectroscopy Method”. (ID: agronomy-2388089). Thank you for giving our opportunity to revise our manuscript again. Thanks for your kind comments on improving our manuscript to be better accepted. We have revised the manuscript carefully and we hope will meet your request and for approval.

The reviewer’s comments are marked in yellow and our responses are marked in green.

Best,

Yongzong Lu （[email protected]） and Yongguang Hu ([email protected])

Q1: Fig.2, is hard to see different plots. Offset each for clarity.

R: Thanks for your kind comments. We have changed the type of line to make the figure 2 more clear.

Figure 2. Absorbance spectra in the band of 0.1–1.6THz of the tea leaf response to different low temperature.

Q2：Fig. 4, 5 could be combined to show the improvement. Also please use the same x scale for Fig. 5b.

R: Thanks for your kind comments. We have revised the figure 4 and 5, improve the quality of the figure to meet the requirement.

Figure 4. AI of THz absorption spectral in different low temperature in the range of 0.1-1.6THz.

(a)

(b)

Figure 5. 2DCOS-THz chart in the range between 0.1THz and 1.6THz: (a) 2DCOS-THz synchronous spectra; (b) The auto-peaks spectra of CI.

The relevant corrections have been made in revised version in the manuscript.

Author Response File: Author Response.pdf