Nutritional Quality, Chemical, and Functional Characteristics of Hemp (Cannabis sativa ssp. sativa) Protein Isolate
, Natalia Vladimirovna Androsova 1, Abduvali Djabarovich Toshev 1 and Hesham Ali El Enshasy 3,4,5
Round 1
Reviewer 1 Report
The article presents the methods of obtaining the isolate as an ecological and industrially cheap method, but this method has been used for many years, there is nothing new in it. Check the bibliography please.
Line 30-31 Why triptophan expressed relative to 100 grams of HPI?
Line 42-43 "Hemp (Cannabis sativa ssp. satvs) is one of the oldest plants cultivated in Asian countryes more than 5 centuries ago" Are you sure? Check the bibliography please.
References to obtaining protein isolates carried out by other research groups should appear (see references 19 and 20)
line 74, 229, 231, 305 references appear in two differents formats
There is no agreement in the way of expressing the results numerically, sometimes decimals are missing (lines 115-118).
Lines 294-297 say that the percentage of carbohydrates is calculated by subtracting the percentage of the other compounds from 100, so it is to be expected that the sum of all the components equals 100, but this is not the case, since in table 1 the sum of the percentages is 118.
line 541 number 45 is missing
Author Response
Response to reviewer #1 comments
First of all, I would like to thank a reviewer for his time and efforts to improve the quality of the manuscript. It is my pleasure to seriously consider the comments point by point in the manuscript. All responses to the comments have been highlighted with yellow color throughout the manuscript.
# The article presents the methods of obtaining the isolate as an ecological and industrially cheap method, but this method has been used for many years, there is nothing new in it. Check the bibliography please.
Response, I completely agree that is not a new method for preparation a plant protein isolate as I have a pervious article on the preparation of protein isolate from Chickpea, lupin, lentil and quinoa (El-Sohaimy, S. A.; Refaay T. M.; Zaytoun M. A. M. Physicochemical and functional properties of quinoa protein isolate. Ann. Agric. Sci., 2015, 60(2), 297–305. http://dx.doi.org/10.1016/j.aoas.2015.10.007, El-Sohaimy, S. A.; Brennan M. A.; Darwish A. M. G.; Brennan C. S. Chickpea Protein Isolation, Characterization and Application in Muffin Enrichment. International Journal of Food Studies, 2021. 10, SI57-SI71. DOI: 10.7455/ijfs/10.SI.2021.a5 and el-Sohaimy S.A, Sitouhy M and El-Masry R. 2007. Isolation and Partial Characterization of Chickpea, lupine and Lentil seed proteins. World Journal of Agricultural Sciences, 3(1), 123-12.9.). So, the current manuscript used this method as a cheapest methodology for industry and ecofriendly for preparing and characterizing the hemp protein isolate to evaluate its quality and suitability for food industry.
# Line 30-31 Why triptophan expressed relative to 100 grams of HPI?
Response: The measuring unit of all amino acids expressed in g/100g not only tryptophan. So, we cannot express all amino acids in g/100g and tryptophan alone in (mg/100g or mg/g).
# Line 42-43 "Hemp (Cannabis sativa ssp. satvs) is one of the oldest plants cultivated in Asian countryes more than 5 centuries ago" Are you sure? Check the bibliography please.
Response: Changed to “Hemp (Cannabis sativa ssp. sativs) is one of the oldest plants cultivated in Asian countries.”
“John M. McPartland,William Hegman, Tengwen Long. Cannabis in Asia: its center of origin and early cultivation, based on a synthesis of subfossil pollen and archaeobotanical studies. Vegetation History and Archaeobotany (2019) 28:691–702 https://doi.org/10.1007/s00334-019-00731-8”
# References to obtaining protein isolates carried out by other research groups should appear (see references 19 and 20)
Response: Other research groups who are previously worked on HPI were already mentioned and discussed throughout the manuscript.
For instance:
(1) Leonard, W.; Zhang, P.; Ying, D.; Fang, Z. Hempseed in food industry: nutritional value, health benefits, and industrial applications. Compr. Rev. Food Sci. Food Saf., 2020, 19 (1), 282-308. DOI: 10.1111/1541-4337.12517
(5) Aiello, G.; Fasoli, E.; Boschin, G.; Lammi, C.; Zanoni, C.; Citterio, A.; Arnoldi, A. Proteomic characterization of hempseed (Cannabis sativa L.). Journal of Proteomics, 2016, 147, 187– 196. https://doi.org/10.1016/j.jprot.2016.05.033/
(6) Malomo, S. A.; Aluko, R. E. A comparative study of the structural and functional properties of isolated hemp seed (Cannabis sativa L.) albumin and globulin fractions. Food Hydrocolloids, 2015, 43, 743-752. https://doi.org/10.1016/j.foodhyd.2014.08.001
(17) Korus, J. M.; Witczak, R.; Ziobro, L. J. Hemp (Cannabis sativa subsp. sativa) flour and protein preparation as natural nutrients and structure forming agents in starch-based gluten free bread. LWT - Food Sci. Technol., 2017, 84.143-150. DOI:10.1016/J.LWT.2017.05.046
# line 74, 229, 231, 305 references appear in two differents formats
Response: The citation of references in the text have been corrected accordingly
# There is no agreement in the way of expressing the results numerically, sometimes decimals are missing (lines 115-118).
Response: The mistakes have been fixed and marked in yellow color
# Lines 294-297 say that the percentage of carbohydrates is calculated by subtracting the percentage of the other compounds from 100, so it is to be expected that the sum of all the components equals 100, but this is not the case, since in table 1 the sum of the percentages is 118.
Response: the values in the table have been corrected accordingly
line 541 number 45 is missing
Response: The reference number (45) has been fixed
Author Response File: 
Author Response.docx
Reviewer 2 Report
I have attached an edited version of the paper. Some major issues. Only one cultivar of hemp was used for the study and there is a great deal of variation amongst varieties, so hard to suggest that the observations from this study are for all hemp.
There is a tendency to overstate how good hemp is rather than just provide the data with a summary at the end.
Comments for author File: 
Comments.pdf
Author Response
Response to reviewer #2 comments
Firstly, I would express my deepest thank to the respected reviewer for the kind effort and time to improve the quality of the manuscript. all comments have been seriously considered point by point throughout the manuscript.
#I have attached an edited version of the paper.
Response: All comments in attached file have been considered accordingly throughout the manuscript and highlighted with a yellow color
# Some major issues. Only one cultivar of hemp was used for the study and there is a great deal of variation amongst varieties, so hard to suggest that the observations from this study are for all hemp.
Response: I am completely agreeing that there is a variation among different varieties. It is according to the differences in the environmental conditions, cultivation conditions, genetic properties of the variety, and others, but in the current study we are focused on the Cannabis sativa ssp. Sativs that cultivating in the Russian territory to evaluate its ability for using in the food applications because our work is already continued for utilizing the protein isolate of this variety for development some protein-rich beverages and bakery products and also developing a protein bars for sport nutrition.
# There is a tendency to overstate how good hemp is rather than just provide the data with a summary at the end.
Response:
The summary has been improved accordingly. In the summary we have just stated that the nutritional and functional properties of the HPI are suitable for food application or not based on the obtained results.
# In the introduction “Hemp seeds contain about 18%-23% protein”, this is not high compared to soybean.
Response: In the current study there is so comparison between Hemp seed and soybean. It is true that is the protein content of hemp seed lower than soybean, but the quality of protein is better that soybean in the term of amino acids composition and the protein digestibility and availability and also in the term of functional properties which important for utilizing it in food application.
Author Response File: Author Response.docx
Reviewer 3 Report
Article Plants-1981767 is of interest.
The work is well drafted and properly structured. Figures and Tables are fine. English needs further improvement, some typos have been highlighted below
I think this work fits the targets of the Journal and therefore I recommend its publication after minor revisions on Plants.
Some revisions for the authors are detailed in the list given below:
First page: remove “.” after the first author
Title: correct “sativs” in “sativa”
Line 10: put “institute” in capital letter
Line 12: “energy” is written incorrectly
Line 16: remove “a” before “rich”
Line 19: the main verb is missing
Line 24-25: write the sentence better
Line 28: remove “was” before “showed”
Line 32: remove “with” before “increasing”
Line 34: put number 4 before “conclusion”
Line 36: write “provides” and not “provide”
Line 38 and 42: correct “sativs” in “sativa”
Line 49: replace “Yso” with “Uso”
Line 56: add “and” between “edestin” and “globulin”
Line 66: add “of” between “shell” and “hemp”
Line 68: replace “format” with “form”
Line 74: remove the reference and leave only the associated number as in the other cases
Line 77: add “is” to “utilized”
Line 92: replace “targeted” with “targeting”
Line 93: put “.” and a space after “proteins”
Line 101: “for” is written in a bad way
Line 103: “from” is written in a bad way
Line 106: add a space between “pH” and “11”
Line 11-112: the main verb is missing
Line 115: add “is” to “presented”
Line 116-117 and in table 1: “crude” is written in a bad way
Line 118-120: there are several mistakes in the sentence, write it better
Line 120: in “the” the initial letter is missing
Line 122: put the subject
Line 124: “plant” is written in a bad way
In paragraph 2.1 the unit of measure of the first values is not specified
Line 136: replace “extraction” with “extracting”
Line 149: replace “affecting” with “affected”
Line 156: replace “are” with “is”
Line 161: remove “which”
Line 172: remove “was” before “showed”
Line 173: remove brackets which include the molecular weights values
Line 176: add “is” before “called” and “a” before “hexamer”
Line 183: add “a” before “widely”
Line 187: remove brackets including values
Line 192: replace “was” with ”were”
Line 196: replace “where” with “with” or add a verb
Line 196-197: remove brackets including values
Line 201-202: write the sentence better
Line 203: remove “was” before “showed”
Line 206: better to write “with tryptophan as the limiting amino acid
Line 209: add “are” before “poor”
Line 210: write better the last part of the sentence (very long sentence and the subject is missing)
Line 212: add “was” to “characterized”
Line 215: “is” is missing twice
Line 217-220: write the sentence better
Line 225-226: the verb is missing in the second part of the sentence
Line 229: put “is” instead of “are”
Line 243: put “were” instead of “was”
Line 248-249: replace “invistigation” with “investigation”
Line 252-254: write the sentence better
Line 265: put “on” in capital letter
Line 266-267: “pH values OF 4, 5 and 6…”
Line 277: remove “is” and replace “emphasize” with “emphasizes”
Line 281: remove brackets to (4); add “linear” between “strong” and “positive”
Line 282: remove the second part of the sentence from “which” to the end of the sentence; put “in” in capital letter
Line 286: no capital letters for “Foaming” and ”Emulsion”
Line 288: no capital letter for “Perfect”
Line 297: remove (1) from the equation
Line 301: A dot is missing
Line 304: replace (2:1v/v) with (2:1, v/v)
Line 305: remove brackets from (Bligh & Dyer, 1959)
Line 315-316: change the sentence. Better writing: “…to extract the protein and pH of the mixture was checked every 15 min to be maintained at pH 11”
Line 317: replace “which contain” with “containing”
Line 320: replace “adjusted” with “adjusting”
Line 322: replace “12000 x g” with “12000xg”
Line 327: remove brackets from (Klompong et al., 2007) and add the number of the reference; add “a” between “for” and “better”
Line 330: replace “7000 Xg” with “7000xg”
Line 338: remove the dot after “min”
Line 341-342: write the sentence better
Line 342: put “the” in capital letter
Line 343: remove the dot after “min”
Line 347-348: write the sentence better
Line 371: replace “Equation (2)” with “equation 2”
Line 371: remove (2) from the equation
Line 378: no capital letter for “Equation”
Line 378: remove (3) from the equation
Line 382: no capital letter for “Equation”
Line 382: remove (4) from the equation
Line 385-386: replace “ml” with “mL”
Line 387: replace “7000 Xg” with “7000xg”
Line 391: no capital letter for “Equation”
Line 392: remove (5) and (6) from the equations
Line 402: the dot at the end of the sentence is missing
Line 405-406: substitute “precipitate” with “precipitating”
Line 409: add space between “phenylalanine” and “+”
Line 413-417: write the sentence better, it’s way too long and complicated
Line 418-419: “beverage industry IS seeking protein-rich products because they ARE preferable for consumers”
Line 419-422: Consequently, the nutritional and functional properties of the hemp seeds have been explored in the current study might be make it MADE applicable not only to milk replacers, but also to other sorts of beverages…”
Some references should be uniformed to the others (date in bold, doi in the same format, journals in italic and abbreviated, homogenous line spacing).
Author Response
Response to reviewer #3 comments
Firstly, I would express my deepest thank to the respected reviewer for the kind effort and time to improve the quality of the manuscript. all comments have been seriously considered point by point throughout the manuscript.
# The work is well drafted and properly structured. Figures and Tables are fine. English needs further improvement, some typos have been highlighted below
Response: Thank you very much for your kind comment, The English and typewriting mistakes have been rechecked throughout the manuscript
# I think this work fits the targets of the Journal and therefore I recommend its publication after minor revisions on Plants.
Response: Thank you, the valuable comments have been seriously considered throughout the manuscript
Some revisions for the authors are detailed in the list given below:
First page: remove “.” after the first author
Removed
Title: correct “sativs” in “sativa”
Corrected
Line 10: put “institute” in capital letter
Corrected
Line 12: “energy” is written incorrectly
Corrected
Line 16: remove “a” before “rich”
Corrected
Line 19: the main verb is missing
Corrected
Line 24-25: write the sentence better
The sentence has been rewritten accordingly
Line 28: remove “was” before “showed”
Removed
Line 32: remove “with” before “increasing”
The sentence “The foaming capacity of HPI was increased with increasing the pH values to reach the maximum value at pH 11 (67.23±3.20 %).” If the word “with has been removed the meaning of the sentence will change”
Line 34: put number 4 before “conclusion”
Corrected
Line 36: write “provides” and not “provide”
Corrected
Line 38 and 42: correct “sativs” in “sativa”
Corrected
Line 49: replace “Yso” with “Uso”
Changed
Line 56: add “and” between “edestin” and “globulin”
The word “and” has been added
Line 66: add “of” between “shell” and “hemp”
Added
Line 68: replace “format” with “form”
Replaced
Line 74: remove the reference and leave only the associated number as in the other cases
Removed
Line 77: add “is” to “utilized”
“is” has been added
Line 92: replace “targeted” with “targeting”
Changed
Line 93: put “.” and a space after “proteins”
Changed
Line 101: “for” is written in a bad way
Corrected
Line 103: “from” is written in a bad way
Corrected
Line 106: add a space between “pH” and “11”
Space has been added
Line 11-112: the main verb is missing
Added
Line 115: add “is” to “presented”
Corrected
Line 116-117 and in table 1: “crude” is written in a bad way
Corrected
Line 118-120: there are several mistakes in the sentence, write it better
Corrected
Line 120: in “the” the initial letter is missing
Corrected
Line 122: put the subject
Corrected
Line 124: “plant” is written in a bad way
Corrected
In paragraph 2.1 the unit of measure of the first values is not specified
Corrected
Line 136: replace “extraction” with “extracting”
Corrected
Line 149: replace “affecting” with “affected”
Corrected
Line 156: replace “are” with “is”
Corrected
Line 161: remove “which”
Removed
Line 172: remove “was” before “showed”
Removed
Line 173: remove brackets which include the molecular weights values
Removed
Line 176: add “is” before “called” and “a” before “hexamer”
Added
Line 183: add “a” before “widely”
Added
Line 187: remove brackets including values
Removed
Line 192: replace “was” with ”were”
Corrected
Line 196: replace “where” with “with” or add a verb
Changed
Line 196-197: remove brackets including values
Removed
Line 201-202: write the sentence better
Re-written
Line 203: remove “was” before “showed”
Removed
Line 206: better to write “with tryptophan as the limiting amino acid
Corrected
Line 209: add “are” before “poor”
Added
Line 210: write better the last part of the sentence (very long sentence and the subject is missing)
Reformatted
Line 212: add “was” to “characterized”
Added
Line 215: “is” is missing twice
Corrected
Line 217-220: write the sentence better
Reformatted
Line 225-226: the verb is missing in the second part of the sentence
Reformatted
Line 229: put “is” instead of “are”
Replaced
Line 243: put “were” instead of “was”
Corrected
Line 248-249: replace “invistigation” with “investigation”
Corrected
Line 252-254: write the sentence better
Reformatted
Line 265: put “on” in capital letter
Corrected
Line 266-267: “pH values OF 4, 5 and 6…”
Corrected
Line 277: remove “is” and replace “emphasize” with “emphasizes”
Corrected
Line 281: remove brackets to (4); add “linear” between “strong” and “positive”
Corrected
Line 282: remove the second part of the sentence from “which” to the end of the sentence; put “in” in capital letter
Corrected
Line 286: no capital letters for “Foaming” and ”Emulsion”
Corrected
Line 288: no capital letter for “Perfect”
Corrected
Line 297: remove (1) from the equation
Removed
Line 301: A dot is missing
Added
Line 304: replace (2:1v/v) with (2:1, v/v)
Corrected
Line 305: remove brackets from (Bligh & Dyer, 1959)
Removed
Line 315-316: change the sentence. Better writing: “…to extract the protein and pH of the mixture was checked every 15 min to be maintained at pH 11”
Changed
Line 317: replace “which contain” with “containing”
Changed
Line 320: replace “adjusted” with “adjusting”
Changed
Line 322: replace “12000 x g” with “12000xg”
Corrected
Line 327: remove brackets from (Klompong et al., 2007) and add the number of the reference; add “a” between “for” and “better”
Corrected
Line 330: replace “7000 Xg” with “7000xg”
Corrected
Line 338: remove the dot after “min”
Moved
Line 341-342: write the sentence better
Reformatted
Line 342: put “the” in capital letter
Corrected
Line 343: remove the dot after “min”
Removed
Line 347-348: write the sentence better
Reformatted
Line 371: replace “Equation (2)” with “equation 2”
Corrected
Line 371: remove (2) from the equation
Corrected
Line 378: no capital letter for “Equation”
Corrected
Line 378: remove (3) from the equation
Corrected
Line 382: no capital letter for “Equation”
Corrected
Line 382: remove (4) from the equation
Corrected
Line 385-386: replace “ml” with “mL”
The mL is correct
Line 387: replace “7000 Xg” with “7000xg”
Corrected
Line 391: no capital letter for “Equation”
Corrected
Line 392: remove (5) and (6) from the equations
Corrected
Line 402: the dot at the end of the sentence is missing
Corrected
Line 405-406: substitute “precipitate” with “precipitating”
Corrected
Line 409: add space between “phenylalanine” and “+”
Added
Line 413-417: write the sentence better, it’s way too long and complicated
The Summary has been reformatted and improved
Line 418-419: “beverage industry IS seeking protein-rich products because they ARE preferable for consumers”
Corrected
Line 419-422: Consequently, the nutritional and functional properties of the hemp seeds have been explored in the current study might be make it MADE applicable not only to milk replacers, but also to other sorts of beverages…”
Corrected
Some references should be uniformed to the others (date in bold, doi in the same format, journals in italic and abbreviated, homogenous line spacing).
All references have been checked
Author Response File: Author Response.docx
Round 2
Reviewer 1 Report
- In the text, all amino acids are expressed as g/100 g of protein, except tryptophan, which is expressed in g/100 g of HPI, see lines 31 and 464, in both cases is 0.22 g/100 g of protein. On the other hand, in table 2, all the amino acids are expressed in g/100 g of protein, including tryptophan, 0.26g/100g of protein. Do you have an explanation for this difference in criteria regarding trytophan, or is it an error.
- Lines 98-99; 106; 109; 113; 247-248; 255 all the words appear together, without spaces, and it is not understood
Author Response
Response to Reviewer#1 comments R2
- In the text, all amino acids are expressed as g/100 g of protein, except tryptophan, which is expressed in g/100 g of HPI, see lines 31 and 464, in both cases is 0.22 g/100 g of protein. On the other hand, in table 2, all the amino acids are expressed in g/100 g of protein, including tryptophan, 0.26g/100g of protein. Do you have an explanation for this difference in criteria regarding trytophan, or is it an error.
Response: Thank you for your valuable comment, All amino acids expressed in g/100 g. The mistake has been corrected in the manuscript to be uniform units
- Lines 98-99; 106; 109; 113; 247-248; 255 all the words appear together, without spaces, and it is not understood
Response: The mistakes have been fixed accordingly. This mistake might be happened in pdf file because in word file everything is OK.
Author Response File: Author Response.docx
Reviewer 2 Report
The authors are to be commended for the improvements made in the article. There are still some minor word usage/grammar issues.
Author Response
The authors are to be commended for the improvements made in the article. There are still some minor word usage/grammar issues.
Response: Thank you for the a valuable comment. The all comments have been deeply considered and the manuscript has been rechecked for grammar and typewriting mistakes.
Author Response File: Author Response.docx
