Peroxidase Activity in Tomato Leaf Cells under Salt Stress Based on Micro-Hyperspectral Imaging Technique
Round 1
Reviewer 1 Report
Formula (2) - need some explanation. VT, W, VS, t - what is it?
Why Fig. 5A gives us 10 feature wavelengths? Need some more explanation, that is not clear, point with 10 features is not an extremum.
The Discussion chapter has links to the tables from almost the beginning of the article. It's not very convenient, maybe it's possible to somehow arrange it so that the tables are next to their discussion.
The discussion section should be expanded and the results compared with other similar research. This is the weakest part of the article.
Author Response
- Formula (2) - need some explanation. VT, W, VS, t - what is it?
Response: Thanks for the your suggestion. According to advice, we revised the explanation for Formula(2) in line 152-154. VT is total volume of extract enzyme solution, W is fresh weight of sample, VS is enzyme liquid volume for determination, and t is reaction time.
- Please add the software used to develop the multivariate models into the materials and methods section.
Response: Thanks for the your suggestion. According to advice, we revised that UnscramblerX10.4 software was used to develop the multivariate models in line 157.
- Why Fig. 5A gives us 10 feature wavelengths? Need some more explanation, that is not clear, point with 10 features is not an extremum.
Response: Thanks for the your suggestion. SPA is used to extract important wavelength. The most important core of SPA is that the algorithm can search for the variable value with the least redundant information from the spectral matrix, so as to minimize the linear correlation between the variables. In Fig.5A, 10 features wavelengths were calculated by Matlab2014a software, which is minimality of the linear correlation between the variables.
- The Discussion chapter has links to the tables from almost the beginning of the article. It's not very convenient, maybe it's possible to somehow arrange it so that the tables are next to their discussion.
Response: Thanks for the your suggestion. The table in the discussion section is mainly used to further explain the phenomena in the table. Therefore, the table is not placed in the discussion section, so as to suit the format requirements of the journal.
5.The discussion section should be expanded and the results compared with other similar research. This is the weakest part of the article.
Response: Thanks for the your suggestion. We revised the discussion section that expanded and added the results compared with other similar research in line 351-382.
Author Response File: 
Author Response.doc
Reviewer 2 Report
Line 12: Peroxidase not POD in first instance.
Line 14: Replace urgent with necessary
Line 18: Replace peroxidase with POD (see Line 12)
Line 23: on "the" interval
Line 25: correlation coefficient has no units and are 3 decimal places too many.
Line 34: SOD not defined CAT not defined "should be catalase (CAT)
Line 35: replace body with plant
Line 36: change was to is "do not change tense mid-sentence"
Line 41: if widely used cite some references please
Line 42: Remove The start with Physical
Lines 46 -50: some references please, especially line 50 after quantitative studies.
Line 51: "a" spectral - add a
Line 56: References please
Line 58: reference please
Line 62: beam(s) area(s) add s to both words
Line 68: Remove in-situ as the work is not done in the field and add ("a" visual)
Lines 74 - 78: I suggest moving from planta to animal analyses is off topic and detracts from the paper (please consider revising).
Lines 80 -82: Animal examples again and definitely not in-situ
Lines 82 -85: animal references reconsider
Line 103: Your work is on tomatoes remove word "meat"
Lines 117 -120: I found this confusing were both fresh and fixed samples used? Please clarify.
Line 120: OCT not defined
Line 133: using "a" Gaio.....
Line 150: change minus to minutes
Line 158: Equation (2) needs terms explained as you did in (1)
Line 161: replace selecting with selection
Line 179: Rc and Rp not defined "correction" and "prediction" need adding
Line 186: SPXY not defined Table 1 has no units. Table 1 does not have data that indicates chlorophyll splits (remedy this paragraph)
Lines 203 - 207: Discussion mingled with the results
Line 230: Table 2 PCS not defined
Line 260: Figures 5 A and B not intuitive as to 10, why not 12??
Line 269: by "the" IVISSA
Line 272: "the" IVISSA and "the SPA"
Lines 330 -331: Some clarification or classing of methodology required as VCPA has the minimum RMSEC but IVISSA has maximum Rc
Line 365: Have the studies been referenced?
Line 388: "the" micro-hyperspectral
Author Response
Line 12: Peroxidase not POD in first instance.
Response: Thanks for the your suggestion. We revised Peroxidase in line 12.
Line 14: Replace urgent with necessary
Response: Thanks for the your suggestion. We revised necessary in line 14.
Line 18: Replace peroxidase with POD (see Line 12)
Response: Thanks for the your suggestion. We revised POD in line 18.
Line 23: on "the" interval
Response: Thanks for the your suggestion. We revised on the interval in line 23.
Line 25: correlation coefficient has no units and are 3 decimal places too many.
Response: Thanks for the your suggestion. We revised that the correlation coefficient and root mean square error of predictionset(RMSEP) are 0.81 and 18.94 U/g•min, respectively in line 25. And 3 decimal places in paper have revised to 2 decimal places.
Line 34: SOD not defined CAT not defined "should be catalase (CAT)
Response: Thanks for the your suggestion. We revised superoxide dismutase (SOD) and catalase(CAT) in line 34.
Line 35: replace body with plant
Response: Thanks for the your suggestion. We revised plant in line 35.
Line 36: change was to is "do not change tense mid-sentence"
Response: Thanks for the your suggestion. We revised ‘is’ in line 37.
Line 41: if widely used cite some references please
Response: Thanks for the your suggestion. We cited 2 references in line 42.
Line 42: Remove The start with Physical
Response: Thanks for the your suggestion. We revised Physical in line 43.
Lines 46 -50: some references please, especially line 50 after quantitative studies.
Response: Thanks for the your suggestion. We cited some references in line 46-50.
Line 51: "a" spectral - add a
Response: Thanks for the your suggestion. We added ‘a’ in line 52.
Line 56: References please
Response: Thanks for the your suggestion. We have cited some references in line 56.
Line 58: reference please
Response: Thanks for the your suggestion. We have cited some references in line 57.
Line 62: beam(s) area(s) add s to both words
Response: Thanks for the your suggestion. We revised light beams in micron areas in line 63.
Line 68: Remove in-situ as the work is not done in the field and add ("a" visual)
Response: Thanks for the your suggestion. We revised in line 68-69.
Lines 74-78: I suggest moving from planta to animal analyses is off topic and detracts from the paper (please consider revising).
Response: Thanks for the your suggestion. We revised detracts about animal analyses from the paper in line 72-79. At present, microscopic hyperspectral imaging technology is mainly used in medical research, but there are few articles about the application of vegetables. Therefore, a large number of medical animal research papers have been cited before.
Lines 80-82: Animal examples again and definitely not in-situ
Response: Thanks for the your suggestion. We revised detracts about animal analyses from the paper in line 72-79.
Lines 82 -85: animal references reconsider
Response: Thanks for the your suggestion. We revised detracts about animal analyses from the paper in line 72-79.
Line 103: Your work is on tomatoes remove word "meat"
Response: Thanks for the your suggestion. I'm sorry for making such a mistake. We revised tomatoes in line 90.
Lines 117 -120: I found this confusing were both fresh and fixed samples used? Please clarify.
Response: Thanks for the your suggestion. We revised that the fresh tissue, transported and stored at low temperature, is fixed by fixation solution for more than 24h in line 105-106.
Line 120: OCT not defined
Response: Thanks for the your suggestion. We revised optimal cutting tem perature(OCT) in line 110.
Line 133: using "a" Gaio.....
Response: Thanks for the your suggestion. We added "a" in line 120.
Line 150: change minus to minutes
Response: Thanks for the your suggestion. We revised minutes in line 137.
Line 158: Equation (2) needs terms explained as you did in (1)
Response: Thanks for the your suggestion. We revised Equation (2) in line 146-147.
Line 161: replace selecting with selection
Response: Thanks for the your suggestion. We revised selection in line 150.
Line 179: Rc and Rp not defined "correction" and "prediction" need adding
Response: Thanks for the your suggestion. We revised Rc and Rp in line 168-170.
Line 186: SPXY not defined Table 1 has no units. Table 1 does not have data that indicates chlorophyll splits (remedy this paragraph)
Response: Thanks for the your suggestion. We revised SPXY in line 177-178, and Table 1 added units.
Lines 203 - 207: Discussion mingled with the results
Response: Thanks for the your suggestion. We have revised results in line 192-201.
Line 230: Table 2 PCS not defined
Response: Thanks for the your suggestion. We have revised PCS in line 169.
Line 260: Figures 5 A and B not intuitive as to 10, why not 12??
Response: Thanks for the your suggestion. We have revised 10 in line 247. The most important core of SPA is that the algorithm can search for the variable value with the least redundant information from the spectral matrix, so as to minimize the linear correlation between the variables. In Fig.5A, 10 features wavelengths were calculated by Matlab2014a software, which is minimality of the linear correlation between the variables.
Line 269: by "the" IVISSA
Response: Thanks for the your suggestion. We have revised the IVISSA in line 257.
Line 272: "the" IVISSA and "the SPA"
Response: Thanks for the your suggestion. We have revised the IVISSA method and the SPA method in line 260.
Lines 330 -331: Some clarification or classing of methodology required as VCPA has the minimum RMSEC but IVISSA has maximum Rc
Response: Thanks for the your suggestion. We have revised the model analysis in line 314-323
Line 365: Have the studies been referenced?
Response: Thanks for the your suggestion. We added some references in line 351.
Line 388: "the" micro-hyperspectral
Response: Thanks for the your suggestion. We revised "the" micro-hyperspectral in line 382.
Author Response File: Author Response.doc
Reviewer 3 Report
After reading the paper, the reviewer has the following comments:
1) Abstract: Please provide the full term of POD before using abbreviation.
2) Line 19: was compared to what? Please clarify.
3) Abstract: Consider to use coefficient of determination instead of the correlation coefficient.
4) Line 24: regarding “roof of error”, do you mean RMSE? Please clarify.
5) Please provide pertinent reference for the argument “Peroxidase (POD) is an enzyme with high activity in plants, and is one of the key enzymes in the enzymatic defense system of plants under stress conditions. It acts synergistically with SOD catalase CAT”.
6) Please provide pertinent reference for the argument “POD is closely related to various important physiological activities of tomato plants. POD can convert toxic hydrogen peroxide (H2O2) into harmless H2O with a series of changes in activity, so it is widely used to evaluate the stress tolerance of plants”.
7) Please provide pertinent reference for the argument “The physical and chemical analysis is often used in traditional catalase detection. The sample pretreatment and experimental methods are cumbersome, and the detection time is long …”
8) Please double-check the typos throughout the paper, e.g. “was introduced[4]”should be “was introduced [4]”.
9) Regarding “At present, hyperspectral imaging technology has been widely used in the macroscopic qualitative and quantitative detection of crops due to its non-invasive, rapid and chemical-free characteristics, but its research in the microscopic field of crop components is limited.” Please provide several related references to support this fact. In addition, papers related to hyperspectral imaging-based plant inspection should be reviewed and included in the section 1.
10) Section 1: Consider to use a table to review the related work on detection of peroxidase activity in plants using micro-hyperspectral imaging if feasible. It can help the reader to quickly grasp the research context.
11) Section 1: related works on the applications of machine learning and especially deep learning (used for micro-hyperspectral image analysis) should be better emphasized.
12) Regarding “ENVI4.8 software was used to extract regions …”, please provide the reference/webpage related to this software package.
13) Why did the authors use ENVI4.8 for extracting the regions of interest?
14) Line 203: Regarding “Under CK and T1 treatments, POD enzyme activity was lower in the upper part than in the middle part and lower part of the leaves, which may be due to the self-regulation mechanism of plant growth affected by the inconsistent duration of stress.” Please provide pertinent references to support this explanation.
15) Regarding “POD enzyme activity of each treatment increased, which was consistent with previous studies.”, please provide the references of the previous studies.
16) Line 388: “Then PLSR and PCR models based …” should be “Subsequently, PLSR and PCR models based …”
17) Section 5: summarize the research findings/achievements in several bullet points.
18) Section 5: provide several sentences to point out the future extensions of the current work.
Author Response
After reading the paper, the reviewer has the following comments:
1) Abstract: Please provide the full term of POD before using abbreviation.
Response: Thanks for the your suggestion. We revised "Peroxidase(POD)" in line 12.
2) Line 19: was compared to what? Please clarify.
Response: Thanks for the your suggestion. We revised that various spectral pre-treatment combined with partial least squares regression models was compared to original spectrum combined with partial least squares regression model in line 18-20.
3) Abstract: Consider to use coefficient of determination instead of the correlation coefficient.
Response: Thanks for the your suggestion. We revised the coefficient of determination and root mean square error of predictionset(RMSEP) are 0.66 and 18.94 U/g•min, respectively in line 25-26.
4) Line 24: regarding “roof of error”, do you mean RMSE? Please clarify.
Response: Thanks for the your suggestion. We revised root mean square error of prediction set(RMSEP) in line 25.
5) Please provide pertinent reference for the argument “Peroxidase (POD) is an enzyme with high activity in plants, and is one of the key enzymes in the enzymatic defense system of plants under stress conditions. It acts synergistically with SOD catalase CAT”.
Response: Thanks for the your suggestion. We added many references in line 34.
6) Please provide pertinent reference for the argument “POD is closely related to various important physiological activities of tomato plants. POD can convert toxic hydrogen peroxide (H2O2) into harmless H2O with a series of changes in activity, so it is widely used to evaluate the stress tolerance of plants”.
Response: Thanks for the your suggestion. We added many references in line 42.
- Please provide pertinent reference for the argument “The physical and chemical analysis is often used in traditional catalase detection. The sample pretreatment and experimental methods are cumbersome, and the detection time is long …”
Response: Thanks for the your suggestion. We added many references in line 45.
8) Please double-check the typos throughout the paper, e.g. “was introduced[4]”should be “was introduced [4]”.
Response: Thanks for the your suggestion. We revised double-check the typos throughout the paper.
9) Regarding “At present, hyperspectral imaging technology has been widely used in the macroscopic qualitative and quantitative detection of crops due to its non-invasive, rapid and chemical-free characteristics, but its research in the microscopic field of crop components is limited.” Please provide several related references to support this fact. In addition, papers related to hyperspectral imaging-based plant inspection should be reviewed and included in the section 1.
Response: Thanks for the your suggestion. We revised several related references in line 56-57.
10) Section 1: Consider to use a table to review the related work on detection of peroxidase activity in plants using micro-hyperspectral imaging if feasible. It can help the reader to quickly grasp the research context.
Response: Thanks for the your suggestion. We revised detracts about animal analyses from the paper in line 72-79. At present, microscopic hyperspectral imaging technology is mainly used in medical research, but there are few articles about the application of plants. Therefore, a large number of medical animal research papers have been cited before.
11) Section 1: related works on the applications of machine learning and especially deep learning (used for micro-hyperspectral image analysis) should be better emphasized.
Response: Thanks for the your suggestion. The paper mainly focus on model building based on micro-hyperspectral image technique. In future, we will apply the deep learning to build model.
12) Regarding “ENVI4.8 software was used to extract regions …”, please provide the reference/webpage related to this software package.
Response: Thanks for the your suggestion. The webpage related to this software package is http://www.downxia.com/downinfo/328985.html.
13) Why did the authors use ENVI4.8 ?
Response: Thanks for the your suggestion. We use ENVI4.8 mainly because the version is easy to learn.
14) Line 203: Regarding “Under CK and T1 treatments, POD enzyme activity was lower in the upper part than in the middle part and lower part of the leaves, which may be due to the self-regulation mechanism of plant growth affected by the inconsistent duration of stress.” Please provide pertinent references to support this explanation.
Response: Thanks for the your suggestion. We revised the paragraph and added pertinent references in line 345-351.
15) Regarding “POD enzyme activity of each treatment increased, which was consistent with previous studies.”, please provide the references of the previous studies.
Response: Thanks for the your suggestion. We revised the paragraph and added pertinent references in line 345-351.
16) Line 388: “Then PLSR and PCR models based …” should be “Subsequently, PLSR and PCR models based …”
Response: Thanks for the your suggestion. We revised several bullet points in line 380-386.
17) Section 5: summarize the research findings/achievements in several bullet points.
Response: Thanks for the your suggestion. We revised Subsequently in line 380-386.
- Section 5: provide several sentences to point out the future extensions of the current work.
Response: Thanks for the your suggestion. We added several sentences to point out the future extensions of the current work in line 388-398.
Author Response File: Author Response.doc
Round 2
Reviewer 1 Report
Comments have been taken into account, the article can be published.
Reviewer 3 Report
I have no further comments.
