Effect of PBAT Biodegradable Mulch Film Extract on Seed Germination and Seedlings Metabolism of Tobacco
Round 1
Reviewer 1 Report
In this manuscript, methyl tert-butyl ether (MTBE) was used to extract organic compounds from PBAT-BMF to evaluate its effects on tobacco seed germination and seedling metabolism. The results showed that six organic compounds were found in the extract of PBAT-BMF, these organic compounds had stress effects on germination, growth of tobacco seed, which significantly changed the metabolism pathway. This study can provide a theoretical basis for the development and application of PBAT-BMF. Overall, this manuscript is suitable for publication in the AGRICULTURE-BASEL with minor revision. Some revision should be considered.
Some detail suggestion
1. Line 23: Abstract (557.27μg g-1) should be 557.27μg g-1
2. Line 32: seriously should be deleted.
3. Introduction: The organic compound in PBAT-BMF should be briefly summarized.
4. Line 44, 50 etc: mulch film and mulching film should be unified.
5. Line 59, 71: metabolites is not appropriate.
6 Line 85: substance should be changed to organic compounds in full manuscript.
7. Line 121: spectrogram should be changed to mass spectrometry.
8. Line 150: cumulative should be changed to total.
9. Line 187: spectrum should be changed to pathway.
10. Line 191: Load figure should be changed to loading scatter plot.
11. VIP of Tab. 2 should be 2 decimal places.
12. Line 275: Which should be changed to which
13. Line 284-285: This sentence should be rewritten.
14. The references should be carefully checked, especially the format. Such as reference 12 and reference 25.
To improve the quality of publication, we recommended to carefully check in full manuscript, such as consistency, reference layout etc.
Author Response
Dear Editor and Reviewers
Thanks for your constructive comments! We deeply appreciate your support on our work. The comments are very helpful! We have carefully modified our manuscript following the reviewers' recommendations and addressed every comment to the best of our knowledge. All changes are highlighted in light blue in the marked copy, and the point-to-point responses are shown below for your easy reference. Following these significant efforts, we would like to thank you again for your consideration to publish our work in Agriculture.
Comments and Suggestions for Authors
In this manuscript, methyl tert-butyl ether (MTBE) was used to extract organic compounds from PBAT-BMF to evaluate its effects on tobacco seed germination and seedling metabolism. The results showed that six organic compounds were found in the extract of PBAT-BMF, these organic compounds had stress effects on germination, growth of tobacco seed, which significantly changed the metabolism pathway. This study can provide a theoretical basis for the development and application of PBAT-BMF. Overall, this manuscript is suitable for publication in the AGRICULTURE-BASEL with minor revision. Some revision should be considered.
Answer: Thank you very much for your recommendation.
Some detail suggestion
- Line 23: Abstract (557.27μg g-1) should be 557.27μg g-1
Answer: Thanks for your advice. We have modified this sentence.
- Line 32: seriously should be deleted.
Answer: Thanks for your advice. " seriously " was deleted.
- Introduction: The organic compound in PBAT-BMF should be briefly summarized.
Answer: Thanks for your advice. The organic compound in PBAT-BMF was briefly summarized. The sentence "In addition, some studies suggest that organic additives of antioxidants, UV stabilizers, fillers, rubbers, lubricants, flame retardants, etc. need to be considered for discussing the ecological toxicity in degradable plastics. These additives contribute to an improvement of the mechanical properties of a polymer." was added.
- Line 44, 50 etc: mulch film and mulching film should be unified.
Answer: Thanks for your advice. "ing" was deleted.
- Line 59, 71: metabolites is not appropriate.
Answer: Thanks for your advice. "metabolites" was changed to "degradation products".
6 Line 85: substance should be changed to organic compounds in full manuscript.
Answer: Thanks for your advice. " substance" had been changed to" organic compounds ".
- Line 121: spectrogram should be changed to mass spectrometry.
Answer: Thanks for your advice. " spectrogram " was changed to" mass spectrometry ".
- Line 150: cumulative should be changed to total.
Answer: Thanks for your advice. " cumulative " was changed to" total ".
- Line 187: spectrum should be changed to pathway.
Answer: Thanks for your advice. " spectrum " was changed to" pathway ".
- Line 191: Load figure should be changed to loading scatter plot.
Answer: Thanks for your advice. " Load figure " was changed to" loading scatter plot ".
- VIP of Tab. 2 should be 2 decimal places.
Answer: Thanks for your advice. We have modified decimal places.
- Line 275: Which should be changed to which
Answer: Thanks for your advice. " Which " was changed to" which ".
- Line 284-285: This sentence should be rewritten.
Answer: Thanks for your advice. This sentence was rewritten as " This study provided a theoretical basis for assessing the ecological safety in the condition of long-term biodegradable mulch film".
- The references should be carefully checked, especially the format. Such as reference 12 and reference 25.
Answer: Thanks for your advice. The references were modified as Journal requirement.
To improve the quality of publication, we recommended to carefully check in full manuscript, such as consistency, reference layout etc.
Answer: Thanks for your advice. We have carefully checked in full manuscript.
Reviewer 2 Report
The manuscript entitled “Effect of PBAT biodegradable mulch film extract on seed germination and seedlings metabolismin of tobacco”, concerns laboratory experiments with the aim of identifying organic compounds from PBAT-BMF and evaluating their effects on tobacco seed germination and seedling metabolism.
GENRAL COMMENTS
The subject of the manuscript falls with the general scope of the Journal and provides new data for the scientific community. The novelty level is high in my opinion, since the paper deals with an innovative topic low studied until now and that will be more and more important in the next years, given the wide diffusion of BDM films. However, the paper shows some gaps that should be addressed before publication, especially concerning the M&Ms (lack of many information) and the level of English language, grammar and style that will benefit strongly if revised by a mother-tongue proof-reader.
SPECIFIF COMMENTS:
- Abstract: It properly summarises the manuscript and highlights the main findings.
- line 20: this is not true because the germinative percentage and germinative energy were not significant, as reported in Figure 1b. The only seemed effects were on time and germinative rate, even if the significance was not reported
- line 46: I suggest providing some short sentences about the biodegradation of BDM films in natural conditions. For instance, recently Fontanazza et al. (Agriculture 2021, 11(8), 782; https://doi.org/10.3390/agriculture11080782) identified from soil particles directly attached to the surface of bioplastics a bacterium responsible for the biodegradation, Pseudomonas putida, and quantified it in the soil through RT-PCR-
- line 68: please avoid the first person
- line 71: has been reported
- lines 73-80: what are the hypotheses of this research? Please add them
- line 83: please add the commercial name of the used PBAT-BMF and the main characteristics (e.g. thickness)
- line 89: please add from where you purchased the reagents
- line 93: at which conditions it was soaked for 12 h? Please add the temperature and the light/darkness
- line 95: which filter did you use? Please add it
- lines 102-107: it is strange that sometimes you wrote at present and other times at past. Please read carefully the manuscript and be homogenous
- line 102: which kind of petri dishes did you use? Please add at least the diameter
- Did you use in total only 6 petri dishes (3 replicates x 2 treatments (T – CK)) for germination experiment? If yes, it is a very low number. If not, please specify in the text
- information about paragraph 2.3 are very low. Any detail was provided about GC-MS analysis, thus making it not repeatable for readers. Please add the complete methodology
- line 116: please change with “the number of germinated seeds”
- line 124: I suppose from each petri dish
- statistical analysis was explained insufficiently. In particular:
· which kind of PCA did you apply (there are many types)?
· which kind of ANOVA did you apply? I think a one-way ANOVA, but please specify in the text
· Were the basic assumptions of ANOVA (homoscedasticity and normality) satisfied before analysis? If yes, which test did you use for them?
· Percentage data need to be transformed before analysis because they not meet a normal distribution. However, non information was provided
· Which was the experiment design of the experiments? Randomize block design or other? Please specify
- line 132: please name it only “Results”
- Figure 1a: please add the error bars for each point and add the significance (if any), because you cannot demonstrate any effect without a statistical analysis. Stated your results, the effects on tobacco seed germination were not significant
- Figure 2: please leave on the y-axis only “cm”. In addition, please specify in the caption the difference between capital and normal letters, because it is not clear as reported
- Please improve the size of Figure 3, it is not readable
- Figure 3a and 3b: please add the percentage of PC1 and PC2
- Figure 3b: please add an appropriate legend about circles and colours
- the discussion is overall appropriate. I only suggest to move also on natural conditions, for instance emphasizing the bridge from biodegradation to possible phytotoxic effects on crops.
- conclusions, line 261: results about seed germination are not significant. The extracts were phytotoxic only against seedling growth.
Author Response
Comments and Suggestions for Authors
The manuscript entitled “Effect of PBAT biodegradable mulch film extract on seed germination and seedlings metabolismin of tobacco”, concerns laboratory experiments with the aim of identifying organic compounds from PBAT-BMF and evaluating their effects on tobacco seed germination and seedling metabolism.
GENRAL COMMENTS
The subject of the manuscript falls with the general scope of the Journal and provides new data for the scientific community. The novelty level is high in my opinion, since the paper deals with an innovative topic low studied until now and that will be more and more important in the next years, given the wide diffusion of BDM films. However, the paper shows some gaps that should be addressed before publication, especially concerning the M&Ms (lack of many information) and the level of English language, grammar and style that will benefit strongly if revised by a mother-tongue proof-reader.
Answer: Thank you very much for your recommendation. We have conducted a professional organization to review the circulation and correctness of the sentences, and correct the errors.
Abstract: It properly summarises the manuscript and highlights the main findings.
Answer: Thank you very much for your recommendation.
line 20: this is not true because the germinative percentage and germinative energy were not significant, as reported in Figure 1b. The only seemed effects were on time and germinative rate, even if the significance was not reported
Answer: Thanks for your advice. This sentence was rewritten as " the germination percentage and germination energy were non-significantly decreased by 2.50% and 1.00%. Whereas,". "also" was deleted.
line 46: I suggest providing some short sentences about the biodegradation of BDM films in natural conditions. For instance, recently Fontanazza et al. (Agriculture 2021, 11(8), 782; https://doi.org/10.3390/agriculture11080782) identified from soil particles directly attached to the surface of bioplastics a bacterium responsible for the biodegradation, Pseudomonas putida, and quantified it in the soil through RT-PCR-
Answer: Thanks for your advice. "The bacterium of Pseudomonas putida identified from soil particles directly attached to the surface of bioplastics were responsible for the biodegradation [8]" and reference 8 (Agriculture 2021, 11(8), 782) were added.
line 68: please avoid the first person
Answer: Thanks for your advice. "my country" was replaced by "China".
line 71: has been reported
Answer: Thanks for your advice. "was" was replaced by "has".
lines 73-80: what are the hypotheses of this research? Please add them
Answer: Thanks for your advice. The sentence "In summary, these data allowed us to come up with the following hypotheses: the organic compounds may have an effect on plant germination and metabolism in the PBAT-BMF. To test this hypothesis, " were added.
line 83: please add the commercial name of the used PBAT-BMF and the main characteristics (e.g. thickness)
Answer: Thanks for your advice. The PBAT-BMF is selected from the products of well-known domestic companies. In order to avoid conflicts of interest, it is not clearly pointed out here. In addition, the sentence " The specifications of the film were 120 cm wide and 0.008 mm thick. " were added.
line 89: please add from where you purchased the reagents
Answer: Thanks for your advice. "purchased from Sigma (St. Louis, MO, USA)." was added.
line 93: at which conditions it was soaked for 12 h? Please add the temperature and the light/darkness
Answer: Thanks for your advice. "at room temperature with dark condition," was added.
line 95: which filter did you use? Please add it
Answer: Thanks for your advice. " with a 0.45 μm nylon membrane and transferred " were added.
lines 102-107: it is strange that sometimes you wrote at present and other times at past. Please read carefully the manuscript and be homogenous
Answer: Thanks for your advice. "is" was replace by "was".
line 102: which kind of petri dishes did you use? Please add at least the diameter
Answer: Thanks for your advice. "(120 mm dia. × 20 mm height)" was added.
Did you use in total only 6 petri dishes (3 replicates x 2 treatments (T – CK)) for germination experiment? If yes, it is a very low number. If not, please specify in the text
Answer: Thanks for your advice. " performed with 6 replicates x 2 treatments (T and CK group)" were specified.
information about paragraph 2.3 are very low. Any detail was provided about GC-MS analysis, thus making it not repeatable for readers. Please add the complete methodology
Answer: Thanks for your advice. The complete methodology was added as follows:
"The PBAT-BMF extract were performed with dispersive liquid–liquid microextraction for acetone/carbon tetrachloride mixture as dispersive solvent and extraction solvent and then determined with gas chromatography-mass spectrometry (GC-MS, Agilent7890A-5975C). HP-5 ms capillary column (60 m × 0.25 mm, 0.25 μm) was used for separation. An injector temperature of 280 °C, split injection, and a split ratio of 5:1 were used. The oven temperature was initially held at 60 °C for 2 min, raised at 15 °C min−1 to 230 °C for 5 min, second rate 5 °C min−1 to 280 °C for 20 min and then the post run was conducted for raised at 3 °C min−1 to 290 °C for 5 min. The constant flow rate of helium carrier gas (99.999%) was maintained at 1 mL min−1. The MS transfer line temperature was 280 °C. The ionization chamber and quadrupole temperatures were 230 °C and 150 °C, respectively. Electron impact mass spectrometric data in the range m/z 50–500 amu were collected, using a scan rate of 3.99 s−1 with an ionization voltage of 70 eV, and the solvent delay was 7 min."
line 116: please change with “the number of germinated seeds”
Answer: Thanks for your advice. "the germinative number" was replaced by " the number of germinated seeds ".
line 124: I suppose from each petri dish
Answer: Thanks for your advice. " from each petri dish " was added.
statistical analysis was explained insufficiently. In particular:
which kind of PCA did you apply (there are many types)?
which kind of ANOVA did you apply? I think a one-way ANOVA, but please specify in the text
Were the basic assumptions of ANOVA (homoscedasticity and normality) satisfied before analysis? If yes, which test did you use for them?
Percentage data need to be transformed before analysis because they not meet a normal distribution. However, non information was provided
Which was the experiment design of the experiments? Randomize block design or other? Please specify
Answer: Thanks for your advice. The statistical analysis was rewritten as follows:
The study was conducted in petri dishes experiments in complete randomized block design with six replicates in two groups. The variation analysis were analyzed by Microsoft Excel 2016 and DPS 17.10 software, principal component analysis (PCA) with euclidean distance as a similarity metric were carried out by SIMCA-P12. One-way ANOVA and multiple comparison performed by Duncan test. The assumption of homoscedasticity was tested using the Levene test. Logarithmic transformation of percentage data helps to meet a normal distribution. Other diagrams were plotted by OriginPro 9.0 software.
line 132: please name it only “Results”
Answer: Thanks for your advice. "and analysis" was deleted.
Figure 1a: please add the error bars for each point and add the significance (if any), because you cannot demonstrate any effect without a statistical analysis. Stated your results, the effects on tobacco seed germination were not significant
Answer: Thanks for your advice. The error bars was added in Figure 1a.
Figure 2: please leave on the y-axis only “cm”. In addition, please specify in the caption the difference between capital and normal letters, because it is not clear as reported
Answer: Thanks for your advice. The annotation of y-axis was added. A and B were P < 0.01 statistical significance, a and b were P < 0.05 statistical significance.
Please improve the size of Figure 3, it is not readable
Answer: Thanks for your advice. The size of Figure 3 was improved.
Figure 3a and 3b: please add the percentage of PC1 and PC2
Answer: Thanks for your advice. The percentage of PC1 and PC2 was added in Figure 3.
Figure 3b: please add an appropriate legend about circles and colours
Answer: Thanks for your advice. X (green) and Y (violet) represent the metabolites and sample center point. X (red) represents a potential biomarker.
the discussion is overall appropriate. I only suggest to move also on natural conditions, for instance emphasizing the bridge from biodegradation to possible phytotoxic effects on crops.
Answer: Thank you very much for your recommendation.
conclusions, line 261: results about seed germination are not significant. The extracts were phytotoxic only against seedling growth.
Answer: Thanks for your advice. "the germination and" was deleted.
References: The order of reference was auto-adjusted in revised manuscript.
Round 2
Reviewer 1 Report
Please check the references carefully
Reviewer 2 Report
The authors addressed all the comments and suggestions previously provided, thus enhancing the overall quality of the manuscript. It is now ready for publication in Agriculture.
