Assessment of an NDL-PCBs Sequestration Strategy in Soil Using Contrasted Carbonaceous Materials through In Vitro and Cucurbita pepo Assays

Round 1

Reviewer 1 Report

The manuscript is focused on the assessment of non-dioxin-like-PolyChloroBiphenyls sequestration strategy in soil using Biochars and activated carbons. Overall, the manuscript is interesting, but some parts need to be re-written to be clearly understood. Some parts of the text are difficult to follow, quite many abbreviations are not explained. I do not like the abstract very much. I suggest to re-write it completely. Since the PCB concentration was measured in plants after harvest (after freezing and extraction, not in intact “live” plants), I would strongly suggest using the term “in planta” not “in vivo”. The manuscript would also benefit from a graphical abstract. All questions and comments follow accordingly to the row number.

Abstract

17 – spelling – planst – plants

21-22 – The whole sentence is unnecessarily too long. “Cucurbita pepo plants were grown for 12 weeks.” is in fact enough.

27 – a space missing “considersuch”

28 – non-dioxin-like-PolyChloroBiphenyls is in a different font

Introduction

CLD, POP, PCB – abbreviations not explained, it may be clear from the context, but it should be explained

Materials and Methods

82 – Explain “aerial parts” of C. pepo – leaves, stems, fruit…?

85 – I would describe the differences among ACs and Biochars here. The link to Table S1 is missing.

91 – Remove the comma.

107 - Table 1 – what is the difference between Control and Standard soil (SS)? It is not clear from the table.

125 – C. pepo should be in italics

126 – Seeds of C. pepo…

133 – Were the plants homogenous? Did they grow to the same size (weight etc.)? You should add more information about the plants. You note (line 196-200) that the plants did not grow properly.

141 + 151 – The paragraphs have the same titles – Statistical analyses. How many samples of soil were tested? The information is missing. The number of plants (line 196-200) should be here as well.

Results

Figure 1 – The legend should contain explanation of abbreviations. The difference between SS and control at least.

Figure 2 – Does DM mean dry mass? Explain in the legend.

231 – The number should be 3.2.1 Environmental availability

Table 2 – There should be decimal points not commas, I suspect.

Figure 4 – What are the values? The title says …. In St Cyprien soil and later … obtained through the in vivo C. pepo assay. The values are from the soil with C. pepo? Re-write the legend more clearly.

Do you have any more information about the plants? Did they vary in the growth parameters or something like? Did you measure pH in soil? It would be better to have more data to support the discussion in the paragraph 4.2.

Author Response

Reviewer #1

We are gratefull for the time spent on your thorough review. You may find attached the response to your comments:

Abstract

17 – spelling – planst – plants

21-22 – The whole sentence is unnecessarily too long. “Cucurbita pepo plants were grown for 12 weeks.” is in fact enough.

27 – a space missing “considersuch”

28 – non-dioxin-like-PolyChloroBiphenyls is in a different font

Please find in the updated manuscript the corrections provided.

Introduction

CLD, POP, PCB – abbreviations not explained, it may be clear from the context, but it should be explained

We added in the document all the definitions of the abbreviations used

Materials and Methods

82 – Explain “aerial parts” of C. pepo – leaves, stems, fruit…?

Indeed, explanations were clearly lacking in the document. Updated document refers now to the different parts of the plants.

85 – I would describe the differences among ACs and Biochars here. The link to Table S1 is missing.

We agree that description of the material are of particular importance to explain the results of the present study. However, as these descriptions are in an article accepted with minor revisions, we can only make them appear briefly, as presented in the table. We updated the reference to make appear the modified status (submitted -> in press) of this cited publication.

We are sorry that you could not find the uploaded supplementary document for your review. We modified Table S1  to add a column to detail the form of the AC or BC.

Table S 1: Physico-chemical characterization of ACs and BCs

Further physical descriptions of the materials are presented elsewhere (El Wanny et al., accepted)

Sample	Furnisher	Origin	Form	Activation	BET surface area (m2.g-1)	Volume mesopore (BJH) (cm3.g-1)
AC1	Roth (ref: 5966.2)	Peat	Granular	Steam	566 ± 5	0.2
AC2	Roth (ref: 5963.2)	Peat	Powder	Phosphoric acid	1142 ± 31	0.6
AC3	Roth (ref: 865.3)	Pine	Powder	Steam	776 ± 21	0.4
BC1	CarboFrance	Oak	Powder	-	28 ± 2	-
BC2	CarboFrance	Oak	Powder	-	281 ± 11	-
BC3	CarboFrance	Knotweed	Powder	-	196± 6	0.03

91 – Remove the comma.

The document was modified accordingly

107 - Table 1 – what is the difference between Control and Standard soil (SS)? It is not clear from the table.

Indeed, the only modification between Control and Standard Soil is the contamination status : controls were not contaminated. The introduction of this modality allow us to consider that background contaminations do not interfer with our present results.

125 – C. pepo should be in italics

The document was modified accordingly

126 – Seeds of C. pepo…

The document was modified accordingly

133 – Were the plants homogenous? Did they grow to the same size (weight etc.)? You should add more information about the plants. You note (line 196-200) that the plants did not grow properly.

No factors limited the initiation and growth of aerial parts during the vegetative phase of the plants. During flowering, a large proportion of the flowers aborted at the end of the experiment, thus limiting optimal fruit development and yield. However the conditions were comparable between groups allowing us to focus on the main scientific question about the extent of the NDL-PCB transfer reduction through this sequestration strategy. 

This paper will allow us to focus further on the improvement of this strategy by assessing the impact on the plant and study closely the transfer mechanisms explaining the differences between both assays.

141 + 151 – The paragraphs have the same titles – Statistical analyses. How many samples of soil were tested? The information is missing. The number of plants (line 196-200) should be here as well.

The document was modified accordingly and presents now the number of repetitions for both assays (n=3 for environmental availability and n=5 for plant assay)

Results

Figure 1 – The legend should contain explanation of abbreviations. The difference between SS and control at least.

We added in the document all the definitions of the abbreviations used in this legend.

Figure 2 – Does DM mean dry mass? Explain in the legend.

Indeed DM stands for Dry Matter and we added this definition in the legend

231 – The number should be 3.2.1 Environmental availability

The document was modified accordingly

Table 2 – There should be decimal points not commas, I suspect.

The document was modified accordingly and all commas were replaced by dots

Figure 4 – What are the values? The title says …. In St Cyprien soil and later … obtained through the in vivo C. pepo assay. The values are from the soil with C. pepo? Re-write the legend more clearly.

The legend is replaced by :  In vivo concentrations of PCB 138, PCB 153 and PCB 180 present in C. pepo aerial parts when grown on the St Cyprien soil.

Do you have any more information about the plants? Did they vary in the growth parameters or something like? Did you measure pH in soil? It would be better to have more data to support the discussion in the paragraph 4.2.

The pH was determined before sowing and no deficiency induced during the growth of the plants was observed. Due to the presence of Calcium Carbonate (known for its buffering capacity) no further assessment of pH was performed. In addition, dynamic variation of pH could not been performed in order to avoid a stress for the plants during their growth. 

Indeed, these preliminary results focus mainly on the sequestration transfer and the impact of NDL-PCBs transfer to the plants. We agree with the reviewer that the developed hypotheses in the discussion section are of particular interest and we will work further to identify the impact of such strategies to the plant growth, and mechanistic considerations to explain the obtained differences. We now clearly indicate this discussion part as hypotheses needing to be addressed.

Reviewer 2 Report

Very interesting topic. I wish all the best in future research. 

Author Response

Reviewer #2

We are gratefull for the time spent on your review and the comments made to this paper. We wish you all the best in your future research.

Reviewer 3 Report

This manuscript is about the study on two different carbonaceous materials to reduce the levels of some very common and persistent PCB congeners (PCB 101, 138, 153 and 180) from the soils to the plants. Both materials, activated carbons and biochars, were evaluated by the addition of a mix of standards of those congeners. After application of the methodology to those samples, clear reductions were detected in the transfer of those POPs to aerial parts of one selected plant (C. pepo). Great results were obtained when the material was activated carbon. 

In my opinion, this work is interesting to apply in areas with soils with notable contamination degrees. However, I consider that the manuscript is not enough clear. I consider that some parts must be extended. Although it is not necessary to develop excessively the materials and methods section, I miss brief information about the analytical procedure as I describe below. I think that this manuscript can be improved with a better structure and explanation. About Supplementary material, I consider it can be included in the normal text because is not too long and a significant part of table S2 is already given in table 2. 

Anyway, I have some comments that I below describe.

 

Orthographic or typewriting doubts

• L. 17. Abstract. ‘Plants’ instead of ‘planst’.
• L. 28. Abstract. Change font on “non-dioxin-like-polychlorobiphenyls”. (Maybe this is something for editorial).
• L. 72. Please, pay attention to the last ‘I’ in ‘… biphenils’. It must be a ‘y’.
• L. 104, 122, 123. Please insert a blank between numbers and units. The same in L. 298 in the species name.
• Figure 5. May authors change the word ‘exsudates’ inside step number 2 into ‘exudates’?
• Please, define some different abbreviations used in the text the first time they appear. In particular, POPs (L. 47), CLD-contaminated (L. 60) and OECD soil (L. 79).

 Materials and methods.

• When authors talk about historically contaminated soils, I consider it necessary to mention their importance or why they are so-called to better understand the manuscript. Do these soils have known and reliable concentrations to be taken as a reference? Since when have been they studying?
• As authors use values from the laboratory test, I think that it is necessary to know where the analysis was carried out and how it was done. I miss a brief paragraph about clean-up, blanks, glassware clean up, solvents and sorbents and quality of them, recoveries. Also, please, add short information about the instrumental analysis and standards (¿native and labelled? )added at the beginning of their treatment and into the extract injected into the equipment.
• About the carbonaceous materials, I haven’t found information about them. The first one, from ROTH is included in a know trademark company, but the second one, from Carbofrance, has no information on the web. I consider it important to know the quality and composition of both materials to know how they can behave about PCB congeners. How they were prepared for the experiments? Are they standardized materials? Were they washed and cleaned up with a protocol of solvents before using? The unique information given is from table S1.
• The text of section 2.2. is finished by a comma. Perhaps a paragraph is missing?
• L. 103. Please, remove ‘cf’ from the parenthesis of Table 1.
• Table 1. Please, put together both lines in the caption.
• Through ignorance, what is the meaning of ‘Diamant cv’ in line 126?

 

Results and discussion

• I would like to know if the authors did not find problems with sulfur. This element is very common and abundant in soils and sediments and makes any analysis very complicated. When it happens separation, identification and peak integration can become impossible, mainly in short retention times.
• L. 163. I consider that PCB 101. 138, 153 and 180 are concentrations. Please, add it.
• Figure 1. I encourage you to enlarge the size of this and the rest of the figures to see clearer its content. In figure 1, please, put together the footnote to avoid an isolated line.  
• L. 287. Authors talk about the granular shape in textural characteristics of carbonaceous materials. This is information that I miss in the materials and methods section.

 

Cites and references

• Please, use abbreviated journal names in all cases. Also, please, remove the issue when is added to the volume.
• Please, change the citation numbers between brackets into the same form: bold or unbold.
• L. 42. Remove text inside the brackets just to show the number of the cite.
• L. 59. Both cites inside the brackets must be shown as their corresponding numbers: 11 and 12.
• L. 302. The cite Hinsinger et al., 2006 is not included in the list of references.
• Please, although they are not mandatory, add the DOI in references 3, 18 and 34.

Author Response

Reviewer #3

We acknowledge the reviewer the reviewer the detailed review, insightful comments and time spent. To answer your thorough review we address a point by point answer to each of your comments.

This manuscript is about the study on two different carbonaceous materials to reduce the levels of some very common and persistent PCB congeners (PCB 101, 138, 153 and 180) from the soils to the plants. Both materials, activated carbons and biochars, were evaluated by the addition of a mix of standards of those congeners. After application of the methodology to those samples, clear reductions were detected in the transfer of those POPs to aerial parts of one selected plant (C. pepo). Great results were obtained when the material was activated carbon. 

In my opinion, this work is interesting to apply in areas with soils with notable contamination degrees. However, I consider that the manuscript is not enough clear. I consider that some parts must be extended. Although it is not necessary to develop excessively the materials and methods section, I miss brief information about the analytical procedure as I describe below. I think that this manuscript can be improved with a better structure and explanation. About Supplementary material, I consider it can be included in the normal text because is not too long and a significant part of table S2 is already given in table 2. 

We totally agree that these results are of interests but the material and method section missed details to clearly state the analytical methodology used. We now address this issue and you will find in the corrected manuscript further details provided especially for this part.

However we can't add table S1 in the regular text of this manuscript as this table is also a part of a paper already published. We updated the citation.

Anyway, I have some comments that I below describe.

 

Orthographic or typewriting doubts

•  
• L. 17. Abstract. ‘Plants’ instead of ‘planst’.
• L. 28. Abstract. Change font on “non-dioxin-like-polychlorobiphenyls”. (Maybe this is something for editorial).
• L. 72. Please, pay attention to the last ‘I’ in ‘… biphenils’. It must be a ‘y’.
• L. 104, 122, 123. Please insert a blank between numbers and units. The same in L. 298 in the species name.
• Figure 5. May authors change the word ‘exsudates’ inside step number 2 into ‘exudates’?
• Please, define some different abbreviations used in the text the first time they appear. In particular, POPs (L. 47), CLD-contaminated (L. 60) and OECD soil (L. 79)

We have now addressed all these typewriting/orthographic mistakes in the corrected document.

 Materials and methods.

•  
• When authors talk about historically contaminated soils, I consider it necessary to mention their importance or why they are so-called to better understand the manuscript. Do these soils have known and reliable concentrations to be taken as a reference? Since when have been they studying?

We are in agreement with the reviewer and now state in the material and methods section : 

"In 22nd of August 2008 a fire broke on this former facility storing large quantities of contaminated oil. This fire event produced a pollution of the agricultural areas nearby resulting in PCB impregnation of soil and subsequently in the destruction of various locally produced foodstuffs, including vegetables. Recent contamination data of the site are provided elsewhere [20] and several studies were performed on various soil samples collected from this site during the past decade [20–22]"

• As authors use values from the laboratory test, I think that it is necessary to know where the analysis was carried out and how it was done. I miss a brief paragraph about clean-up, blanks, glassware clean up, solvents and sorbents and quality of them, recoveries. Also, please, add short information about the instrumental analysis and standards (¿native and labelled? )added at the beginning of their treatment and into the extract injected into the equipment.

Indeed, the material and methods clearly missed further details to provide insigth of the analytical steps and the methodology used. Laboratoire d'Analyses des Sols d'Arras performed all the analyses of NDL-PCBs using normalised and/or accreditated methodologies. We now inserted further description about the quality of this analytical methods and we thoroughly modified this section : 

"2.6. Analytical processes

All chemicals used during the analytical processes were of Pesticide Residues grade and glassware equipment was cleaned with an alkaline detergent, (RBS T 105, Carl Roth GmbH, Karlsruhe, Germany).

2.6.1 Extraction processes of plant samples

Biological matrices were extracted using Pressurized-Liquid Extraction (PLE) and purified using acidified silica columns as previously described [20]. 

2.6.2 Purification step

Potential chromatographic interferences were removed from extracts using Multi-layer silica column (28397-U, Merck KGaA, Darmstadt, Germany). The silver nitrate treated layer removes sulphur-containing compounds; whilst two sulphuric acid treated layers oxidise sample lipids and remove any basic analytes. The potassium hydroxide treated layer removes any acidic sample components. NDL-PCB pass through the silica column unretained. Each extract was evaporated until dryness and dissolved in nonane.

2.6.3 Quantitation processes 

Quantification of NDL-PCBs was performed the extracts of the soil environmental availability and plants using a Gas Chromatography - High Resolution Mass Spectrometry (GC-HRMS) device in Laboratoire d’Analyse des Sols (Arras, Hauts de France, France). Quantification was performed by selecting the two most abundant molecular ions for each organic compound, by isotopic dilution and at a resolution close to 10,000. The determination was performed in SIR (Single Ion Recording) mode and ionization by electron impact in low energy positive mode (35.5 eV) and with a trap current between 400 and 600 µA. Program of the column (DB-5MS 60m 250µm internal diameter, 0.25µm active phase thickness, Agilent J&W, Santa Clara, USA) consisted of a steady temperature gradient from 100°C to 325°C (44 min).

2.7. Quality control of the analytical processes

Quantification was performed using isotopic dilution by adding 13C internal standards (EC9605-SS, Wellington Laboratories, Canada) as described in Environment Canada Method 1/RM/31. Ratio of the molecular ions area of the parent compounds over internal standards of the same chlorine number allowed the quantification after calibration (Regression coefficient R² >0.99). In addition, perfluorokerosene (Lock mass) was added allowing mass calibration of the mass spectrometer during the whole analysis. Recoveries of the whole analytical steps above 70% were obtained throughout analytical runs using recovery standards (EC9605-RS, Wellington Laboratories, Canada). In each run, chromatographic and method blanks were introduced showing no cross-contamination (all < LOQs). A quality control using an independent standard solution was introduced showing a <20% deviation. Limits of quantification for 2g of soil or plants are 0.2 µg.kg-1 for each NDL-PCB. Each steps of the analytical process were performed as documented in the COFRAC quality accreditation of LAS laboratory for the NDL-PCBs quantification of soil samples."

• About the carbonaceous materials, I haven’t found information about them. The first one, from ROTH is included in a know trademark company, but the second one, from Carbofrance, has no information on the web. I consider it important to know the quality and composition of both materials to know how they can behave about PCB congeners. How they were prepared for the experiments? Are they standardized materials? Were they washed and cleaned up with a protocol of solvents before using? The unique information given is from table S1.

Indeed, we have not introduced in this manuscript any production details concerning the matrices. Carbofrance is a leading producer of charcoals based in Montier sur Saulx in France and was involved as a partner of the project PIEGEACHLOR. In this frame, different matrices were produced as described previously in Yehya 2018 using different type of wood. The following further descriptions were added to the manuscript : 

"Biochars were produced by pyrolysis using the industrial-scale ovens of CARBOFRANCE (Montiers-sur-Saulx, Lorraine, France). Oak stems originating from various locations of the East part of France, to ensure comparability with the raw material of this industry was used. Oven were set ot perform a pyrolysis at 500°C or 700°C. After the pyrolysis process, biochars samples were ground and sieved until <500 μm before their use."

• The text of section 2.2. is finished by a comma. Perhaps a paragraph is missing?
• L. 103. Please, remove ‘cf’ from the parenthesis of Table 1.
• Table 1. Please, put together both lines in the caption.

We have now addressed all these typewritting/orthographic mistakes in the corrected document.

• Through ignorance, what is the meaning of ‘Diamant cv’ in line 126?

As this abbreviation is common in the botanic field but not totally in environmental science, we choose to replace it by the non-abbreviated form : cultivar.

Results and discussion

• I would like to know if the authors did not find problems with sulfur. This element is very common and abundant in soils and sediments and makes any analysis very complicated. When it happens separation, identification and peak integration can become impossible, mainly in short retention times.

Indeed, sulfur is a main interferent in such analyses. To perform accurate quantification, a purification step was performed. We introduced further description of this purification involving specific phase to limit such interferent. 

• L. 163. I consider that PCB 101. 138, 153 and 180 are concentrations. Please, add it.

Mention was added in the caption.

• Figure 1. I encourage you to enlarge the size of this and the rest of the figures to see clearer its content. In figure 1, please, put together the footnote to avoid an isolated line. 

We have enlarged the size of the figures to improve its readability

• L. 287. Authors talk about the granular shape in textural characteristics of carbonaceous materials. This is information that I miss in the materials and methods section.

We have now introduced a form section in the Table S1.

Cites and references

• Please, use abbreviated journal names in all cases. Also, please, remove the issue when is added to the volume.

We checked the citation to use the abbreviated journal names

• Please, change the citation numbers between brackets into the same form: bold or unbold.

We have addressed this formatting issue.

• L. 42. Remove text inside the brackets just to show the number of the cite.

We have changed this citation to fulfill the guidelines of the journal

• L. 59. Both cites inside the brackets must be shown as their corresponding numbers: 11 and 12.

We have changed this citation to fulfill the guidelines of the journal

• L. 302. The cite Hinsinger et al., 2006 is not included in the list of references.

We have changed this citation to fulfill the guidelines of the journal

• Please, although they are not mandatory, add the DOI in references 3, 18 and 34.

We have introduced now the doi in all references that have one.

Round 2

Reviewer 3 Report

I would like to thank the authors for the effort to get this revised manuscript. I still have some comments about several parts of the text.

 

• I think that all names must be always written in the same form. I am talking about the name of PCBs. In my opinion, the right one is non-dioxin-like polychlorinated biphenyls (NDL-PCBs). So, change it in the abstract and other parts of the text, as in line 72.
• I also don’t consider it necessary to use capital letters to talk of POPs. So, I consider that persistent organic pollutants (POPs) are the best form (L. 47). The same for chlordecone (CD) in line 60.
• In the caption of figure 1, please, lines 125 and 126 must be put together in just one line.
• Section 2.6. Are all solvents, reagents and the alkaline detergent from Carlo Roth?

Have authors used any protocol for cleaning up all lab material using different solvents o heating them in an oven above 400 ºC or even higher? PCBs usually are adsorbed to glass and need to be strongly treated to remove them, what would be cross-contamination and signals of those compounds could interfere in the chromatographic process before analysis by MS. 

• Section 2.6.3. L. 171. I think that the sentence needs an ‘on’ between ‘performed’ and ‘ the extracts’.

1. 174. Which GC-MS did you use? Please, add the trademark, model and country of the company.
2. 181. Please, insert blanks between numbers and units in the description of the column.

Related to the temperature of analysis in the GC, I think that it is a bit rare. Have you used a ramp with a rate of 5 ºC/min? Usually, it’s common for a strong ramp for the first minutes and a second one or even a third one to reach the temperatures to let the higher chlorinated congeners go out the column. In this way, I think it wouldn’t be necessary to go to so high temperatures to extend the life of the column. DB-5MS has a limit of 350 ºC working with temperature ramps.

• Section 2.7. L. 190. Better to write ‘lock mass’, with an initial lower case letter.

1. 195. What is the independent standard solution? Which concentration?
2. 196-199. Please, unify criteria: quantification or quantitation wherever you must use it.

• Figure 2. L. 247. As in some former comments, I consider it better to write polychlorinated biphenyls (PCBs), with an initial lower case letter.

 

 

Author Response

Open Review

(x) I would not like to sign my review report
( ) I would like to sign my review report

English language and style

( ) Extensive editing of English language and style required
( ) Moderate English changes required
( ) English language and style are fine/minor spell check required
(x) I don't feel qualified to judge about the English language and style

 

	Yes	Can be improved	Must be improved	Not applicable
Does the introduction provide sufficient background and include all relevant references?	(x)	( )	( )	( )
Is the research design appropriate?	( )	(x)	( )	( )
Are the methods adequately described?	( )	(x)	( )	( )
Are the results clearly presented?	( )	(x)	( )	( )
Are the conclusions supported by the results?	( )	(x)	( )	( )

Comments and Suggestions for Authors

I would like to thank the authors for the effort to get this revised manuscript. I still have some comments about several parts of the text.

 We would like to thanks again the reviewer for his insightful comments

• I think that all names must be always written in the same form. I am talking about the name of PCBs. In my opinion, the right one is non-dioxin-like polychlorinated biphenyls (NDL-PCBs). So, change it in the abstract and other parts of the text, as in line 72.

 

All the non-abbreviated occurrences of PCBs were replaced by the correct form, as suggested by the reviewer.

 

• I also don’t consider it necessary to use capital letters to talk of POPs. So, I consider that persistent organic pollutants (POPs) are the best form (L. 47). The same for chlordecone (CD) in line 60.

 

All the non-abbreviated occurrences of POPs and chlordecone are now in regular

 

• In the caption of figure 1, please, lines 125 and 126 must be put together in just one line.

 

We had replaced it in the last version of manuscript, however Word application did not make appear such type of modification in the track version of this manuscript.

 

• Section 2.6. Are all solvents, reagents and the alkaline detergent from Carlo Roth?

Have authors used any protocol for cleaning up all lab material using different solvents o heating them in an oven above 400 ºC or even higher? PCBs usually are adsorbed to glass and need to be strongly treated to remove them, what would be cross-contamination and signals of those compounds could interfere in the chromatographic process before analysis by MS. 

 

The glassware equipment was cleaned with an alkaline detergent, RBS T 105, recommended for cleaning and efficient to remove organic residues, fats, oils, etc. It rinses easily without leaving any interfering residues. All glass materials are put for one night in a bath containing RBS T 105 diluted in demineralized water. The glass materials are then rinsed with demineralized water and dried. All glass materials is rinsed with pure acetone and dried before being used for analysis. The method blanks permit to verify the absence of pollution.

 

• Section 2.6.3. L. 171. I think that the sentence needs an ‘on’ between ‘performed’ and ‘ the extracts’.

We have now corrected this sentence accordingly

1. 174. Which GC-MS did you use? Please, add the trademark, model and country of the company.

The revised manuscript provides now the details concerning the model, trademark and country of the analytical instruments.

 

1. 181. Please, insert blanks between numbers and units in the description of the column.

This typographic issue has been addressed

 

Related to the temperature of analysis in the GC, I think that it is a bit rare. Have you used a ramp with a rate of 5 ºC/min? Usually, it’s common for a strong ramp for the first minutes and a second one or even a third one to reach the temperatures to let the higher chlorinated congeners go out the column. In this way, I think it wouldn’t be necessary to go to so high temperatures to extend the life of the column. DB-5MS has a limit of 350 ºC working with temperature ramps.

 

Indeed, we performed according modification in the revised manuscript.

The furnace temperature program is :

- step of 1 min at 100°C

       - ramp up to 200°C at 40°C/min

       - step of 6 min at 200°C                                                                          

- ramp up to 235°C at 3°C/min

  - step of 10 min at 235°C

- ramp up to 315°C at 8°C/min

- step of 9 min at 315°C

Total time run for analysis : 50.17 min

Temperature of the transfer line between GC and HRMS at 250°C.

 

 

• Section 2.7. L. 190. Better to write ‘lock mass’, with an initial lower case letter.
• This typographic issue has been addressed

1. 195. What is the independent standard solution? Which concentration?

External standard solution ECPCS3 at 200 µg/L from a different batch of the calibration range.

 

196-199. Please, unify criteria: quantification or quantitation wherever you must use it.

Please find in the revised manuscript the use of quantitation instead of quantification

 

Figure 2. L. 247. As in some former comments, I consider it better to write polychlorinated biphenyls (PCBs), with an initial lower case letter.

• This typographic issue has been addressed

Author Response File: Author Response.docx