Red Oak (Quercus rubra L.) Fruits as Potential Alternative for Cocoa Powder: Optimization of Roasting Conditions, Antioxidant, and Biological Properties

Round 1

Reviewer 1 Report

Introduction:

It should be better explained if the antiproliferative effect which is said to exist in vitro could exist also in vivo, and the same for the antibacterial effect since the author is talking about a product to be eaten. 

• Line 64: There are a list of countries and it is included Sardinia which is no a country.
• Line 65. A dot should be written between America and Oak. 
• Line 66 cake is redundant since it is mentioned in the line before in a lis of uses of acorns.
• Line 81-82, It is said the there is an antibacterial effect against some bacteria species but a fungal species is included, Candida albicans. It should be expressed in a different way if the acorns also have the same effect against the fungus.

Statistical Analysis 

Authors said all the analysis were done in duplicate, and they should have been done at least in triplicate to have reliable results. Why didn't you do a triplicate at least?

The main objetive seem to be the potential use of acorn powder as a substitute of cocoa powder and the results seem to conclude that it is possible based on hedonic test, but there are some test mixed like antibacterial activity, the polyphenols content or the anti proliferative properties that are not really related to the potential use of acorn powder since they are tested in vitro and it is not said what happen in vivo in addition to the fact that all the analysis were done in duplicate.

Antibacterial properties seem to be only for one bacteria species, which is not enough to affirm that the extract has antibacterial effect.

Is there any "in vivo" studies to know if the "in vitro" properties work as well in vivo?, since the acorn powder has to be eaten and pass through the digestive system.

 

Author Response

Subject: Submission of revised manuscript No. Forests: 1299438

 

Dear Special Issue Editor, Dr. Apostolos P. Kyriazopoulos

Dear Reviewers,

 

Thank you for considering our manuscript, “Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties” submitted for publication in your esteemed Journal Forests MDPI, Special Issue "Non-wood Forest Products".

As suggested by your editorial office, the authors have responded point by point to all the concerns raised by the reviewers. Also enclosed is a detailed description of the revisions made to the manuscript. We have highlighted in the text the changes we made.

We thank the reviewers for their constructive suggestions. In response to their comments, our manuscript has been improved.

Editorial comments:

Reviewer 1

Thank you for all your encouraging comments.

Comment: Introduction:

It should be better explained if the antiproliferative effect which is said to exist in vitro could exist also in vivo, and the same for the antibacterial effect since the author is talking about a product to be eaten. 

Authors response: To the best of our knowledge, to date, there is no supporting data to make any inferences in the introduction about any in vivo antiproliferative effect of red oak fruits. We present in the Results and discussion section of the manuscript (L367-371) data on the antiproliferative activities of other Quercus species. However, based on the reviewer recommendation, we introduced the following sentence in the manuscript “While it is known that natural sources are of paramount importance summarizing 60% of the currently used anticancer agents (Samarghandian et al., 2010), and the in vitro results are promising, in vivo studies are necessary to ultimately confirm a biological effect.” – lines 372-375. Moreover, this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material (most of the collected acorn material was consumed in preliminary / prefeasibility studies for roasting trials, product development, sensory analysis, etc.) and the current work aimed to develop the acorn powder-based product, the in vitro antioxidant and biological properties of acorns being studied. Furthermore, as the reviewer mentioned, for the future work we intend to perform in vivo studies as well, interdisciplinary research with a research group from University of Medicine being sought.

Comment: Line 64: There are a list of countries and it is included Sardinia which is no a country.

Authors response: corrected

Comment: Line 65. A dot should be written between America and Oak. 

Authors response: corrected

Comment: Line 66 cake is redundant since it is mentioned in the line before in a lis of uses of acorns.

Authors response: corrected

Comment: Line 81-82, It is said the there is an antibacterial effect against some bacteria species but a fungal species is included, Candida albicans. It should be expressed in a different way if the acorns also have the same effect against the fungus.

Authors response: corrected

Comment: Statistical Analysis 

Authors said all the analysis were done in duplicate, and they should have been done at least in triplicate to have reliable results. Why didn't you do a triplicate at least?

Authors response: Dear reviewer, the analysis in this research were done in duplicate due to the lack of materials. As previously mentioned, this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material, most of the collected acorn material being consumed in preliminary / prefeasibility studies for roasting trials, product development, sensory analysis, etc.

Comment: The main objetive seem to be the potential use of acorn powder as a substitute of cocoa powder and the results seem to conclude that it is possible based on hedonic test, but there are some test mixed like antibacterial activity, the polyphenols content or the anti proliferative properties that are not really related to the potential use of acorn powder since they are tested in vitro and it is not said what happen in vivo in addition to the fact that all the analysis were done in duplicate.

Authors response: This study was aimed at investigating the acorn derived powder as an alternative for cocoa powder in order to obtain a chocolate that does not contain stimulants of the nervous system. Based on the hedonic test, we showed the degree of acceptability of the new acorn powder based product obtained. Besides the sensory properties of the new confectionery product obtained, it was aimed to perform a preliminary study of the biological and antioxidant properties of the proposed key ingredient – roasted acorn powder in order to show its potential advantages as a future food ingredient; it does not contain stimuli of the nervous system / successfully replacing cocoa and carob powder, has a high content of polyphenols, has the ability to inhibit the growth of Gram-positive bacterium (Bacillus cereus) and have the ability to reduce the cell viability of the HFL-1 (human fetal lung fibroblast) and DLD-1 (colorectal adenocarcinoma) lines.

Comment: Antibacterial properties seem to be only for one bacteria species, which is not enough to affirm that the extract has antibacterial effect.

Authors response: The research studied the antimicrobial effects of aqueous extracts for Salmonella Enteritidis, Salmonella Typhimurium, Staphylococcus aureus, Bacillus cereus. Following the analyzes, it was concluded that aqueous acorn extracts have the ability to inhibit growth of Bacillus cereus. Throughout the manuscript it is specified that it has a limited antimicrobial effect, that is specific for Bacillus cereus.

Comment: Is there any "in vivo" studies to know if the "in vitro" properties work as well in vivo?, since the acorn powder has to be eaten and pass through the digestive system.

Authors response: We agree the reviewer recommendation about the necessity of studying the in vivo, besides the in vitro properties, so for the future work we intend to perform in vivo studies as well, interdisciplinary research with a research group from University of Medicine being sought. As mentioned previously this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material and the current work aimed to develop the acorn powder-based product, in addition, the in vitro antioxidant and biological properties of acorn powder as a key raw material were also studied. We did not find any  studies for extrapolating the in vitro studies. However, there is a pattern of new food products development and characterization studies / published papers, where the techno-functional properties of some acorn based food products was performed: acorn bread [Kim&Joo, 2019; Shishehbor et al., 2020], muffins with added acorn jelly powder and acorn ethanol extract powder [Kim et al., 2012], gluten – free biscuits made from corn – acorn flour [Korus et al., 2017], biscuits with acorn flour [Pasqualone et al., 2019; Amina& Djamel, 2018; Šálková et al., 2011; Parsaei et al., 2018]

Kim, J.-M., & Joo, J-I. (2019). Quality characteristics of acorn bread added with milk. Journal of the Korean Society of Food Culture, 34(3), 343–352. https://doi.org/10.7318/KJFC/2019.34.3.343

Kim, S. H., Lee,W. K., Choi, C. S., & Cho, S. M. (2012). Quality characteristics of muffins with added acorn jelly powder and acorn ethanol extract powder. Journal of the Korean Society of Food Science and Nutrition, 41(3), 369–375.

Korus, A., Gumul, D., Krystyjan, M., Juszczak, L., & Korus, J. (2017). Evaluation of the quality, nutritional value and antioxidant activity of gluten – free biscuits made from corn – acorn flour or corn – hemp flour biscuits. European Food Research and Technology, 243(8), 1429–1438. https://doi.org/10.1007/.s00217-017-2853-y

Pasqualone, A., Makhlouf, F. Z., Barkat, M., Difonzo, G., Summo, C., Squeo, G., & Caponio, F. (2019). Effect of acorn flour on the physico-chemical and sensory properties of biscuits. Heliyon, 5(8), e02242. https://doi.org/10.1016/j.heliyon.2019.e02242

Amina, M., & Djamel, F. (2018). Effect of fermentation and germination treatments on physicochemical and sensory properties of enriched biscuits with acorn flour. Annals. Food Science and Technology, 19(4), 667–674.

Šálková, T., Divišová, M., Kadochová, Š., Delawská, K., Kadlčková, E., Němečková, L., ... Žemličková, A. (2011). Acorns as a food resource. An experiment with acorn preparation. Interdisciplinaria Archaelogica Natural Sciences in Archeology, 2(2), 133–141. https://doi.org/10.24916/iansa.2011.2.6

Shishehbor, F., Salimi, Z., Veissi, M., Malehi, A. S., Shiri-Nasab, M., &Helli, B. (2020). Effect of oak flour on glycemic index and satiety index of white bread. Iranian Red Crescent Medical Journal, 22(1), 1–7. https://doi.org/10.5812/ircmj.95552

Parsaei,M., Goli,M., & Abbasi, H. (2018). Oak flour as a replacement of wheat and corn flour to improve biscuit antioxidant activity. Food Science & Nutrition, 6(2), 253–258. https://doi.org/10.1002/ fsn3.524

 

Reviewer 2

Comment: The article "Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties" presented by Romina Alina (Vlaic) Marc and co-authors is interesting and useful to the community studying food chemistry and nutrients.  In my opinion, this article should be accepted after minor revision.

Authors started with a good introduction to their research with sufficient literature research.

Their methods are well described. They succeed to present their results clearly.

However, there are some minor issues that require attention:

Authors response: Thank you for all encouraging comments.

Comment: Abstract, Page 1, line 29:  authors may delete “in conclusion” if they wish, and start the sentence with “This study suggested…”

Authors response: corrected

Comment: Page 1, line 39:  authors may introduce what LDL stands for.

Authors response: corrected

Comment: Page 3, line 127: “A Na2CO 7.5%”, should be replaced with “Na2CO3”. (sodium bicarbonates formula should be written correctly. It is better to remove “A” at the beginning of the sentence.

Authors response: corrected

Comment: Page 6, in line 234, and in other lines and tables in the articles: authors did not write “°C” in a correct way. Sometimes they did not introduce any spaces “X°C”, sometimes they introduce a space between the value and °C, sometimes they add a space between ° and C, and sometimes they add 2 spaces. Please notice that °C are connected. Authors can write X°C (without space) or X °C (with a space) but should be homogenous within the text.

Authors response: corrected

Comment: Page 8, line 296: rations should be written in a similar way. Either “X:Y” or “X : Y”.

Authors response: corrected

 

Reviewer 3

Manuscript entitled " Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties " presented to evaluate its sensory and nutritional properties as a potential alternative for cocoa powder in the manufacture of chocolate.

Authors response: Thank you for all encouraging comments.

Comment: Introduction section

In this paragraph it is necessary that the authors explain the advantage and disadvantage of optimization of roasting conditions with different time and temperature intervals. Please add references.

Please add and explain the effect of roasting time and roasting temperature on the color of cocoa powder and acorn powder for sensory properties. These effects are pivotal to optimize the roasting operations of cocoa and acorn. Please add references.

Materials and Methods section

Please clearly elucidate the operation weight of the sample when the roasting performance of the acorn is evaluated.

 

Authors response: the selection of parameters (temperature & time) for acorns’ roasting operation was done in an empirical way, while no other similar studies were reported in literature for acorn or other forest seeds. Moreover, the present study might be considered a PIONEERING STUDY for acorn roasting aiming to replace cocoa powder in confectionery products. Consequently, as presented in Table 1, empirically selected temperatures of 135 and 160 °C were studied, the organoleptic profile (color / flavor) being monitored for 90 minutes, samples being analyzed in 10 / 15 min intervals. Based on the same rationale, the higher temperatures (180, 200 and 220 °C) were also selected empirically; however, the sampling time intervals was decreased to 5 min (table 2 and 3) and the total roasting time to 25 min (after this time, material burning was detected). Moreover, the Results and Discussion section 3.1 Optimization of roasting for acorn powder production, presents the obtained results (clear & organized tables with roasting time & temperature parameters / organoleptic profiles) and critically discuss them.

Comment: L 97-98 Please explain deeply the operation of conditioning operation.

Authors response: corrected

Comment: L 99 Please clearly elucidate the condition of humidity and room temperature when roasting procedure was performed.

Authors response: corrected.

Comment: Please add the quantitative measures were performed by means of the International Commission on Illumination (CIE) L*a*b* when the roasting performance of acorn is evaluated.

Results and Discussion

Please clearly elucidate the values of L*, a*, b*, and the color difference ΔE when the roasting performance of acorn is evaluated, before and after roasting operation at 220 oC/15 min.

Please compare the values of L*, a*, b*, and the total color difference ΔE when roasting operation of the acorn at 220 °C/15min with commercial cocoa powder.

Authors response: Dear reviewer, in the experimental design of this research we did not include the instrumental color analysis due to the lack of the available equipment. However, the color was assessed during the organoleptic analysis, the results being presented in Table 1, 2 and 3. Thank you for this suggestion, in the future research we will include this analysis in the experimental design, a colorimeter being recently bought in our laboratory; however, even now the equipment is available in our laboratory, we are not able to perform the instrumental color analysis, while all collected acorn material was consumed.

Thank you once again for your valuable comments. The authors are available if there are any further queries.

--

 

Best regards,

Marc (Vlaic) Romina Alina et al.

Author Response File: Author Response.docx

Reviewer 2 Report

The article "Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties" presented by Romina Alina (Vlaic) Marc and co-authors is interesting and useful to the community studying food chemistry and nutrients.  In my opinion, this article should be accepted after minor revision.

Authors started with a good introduction to their research with sufficient literature research.

Their methods are well described. They succeed to present their results clearly.

However, there are some minor issues that require attention:

Abstract, Page 1, line 29:  authors may delete “in conclusion” if they wish, and start the sentence with “This study suggested…”

Page 1, line 39:  authors may introduce what LDL stands for.

Page 3, line 127: “A Na₂CO 7.5%”, should be replaced with “Na₂CO₃”. (sodium bicarbonates formula should be written correctly. It is better to remove “A” at the beginning of the sentence.

Page 6, in line 234, and in other lines and tables in the articles: authors did not write “°C” in a correct way. Sometimes they did not introduce any spaces “X°C”, sometimes they introduce a space between the value and °C, sometimes they add a space between ° and C, and sometimes they add 2 spaces. Please notice that °C are connected. Authors can write X°C (without space) or X °C (with a space) but should be homogenous within the text.

Page 8, line 296: rations should be written in a similar way. Either “X:Y” or “X : Y”.

Author Response

Subject: Submission of revised manuscript No. Forests: 1299438

 

Dear Special Issue Editor, Dr. Apostolos P. Kyriazopoulos

Dear Reviewers,

 

Thank you for considering our manuscript, “Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties” submitted for publication in your esteemed Journal Forests MDPI, Special Issue "Non-wood Forest Products".

As suggested by your editorial office, the authors have responded point by point to all the concerns raised by the reviewers. Also enclosed is a detailed description of the revisions made to the manuscript. We have highlighted in the text the changes we made.

We thank the reviewers for their constructive suggestions. In response to their comments, our manuscript has been improved.

Editorial comments:

Reviewer 1

Thank you for all your encouraging comments.

Comment: Introduction:

It should be better explained if the antiproliferative effect which is said to exist in vitro could exist also in vivo, and the same for the antibacterial effect since the author is talking about a product to be eaten. 

Authors response: To the best of our knowledge, to date, there is no supporting data to make any inferences in the introduction about any in vivo antiproliferative effect of red oak fruits. We present in the Results and discussion section of the manuscript (L367-371) data on the antiproliferative activities of other Quercus species. However, based on the reviewer recommendation, we introduced the following sentence in the manuscript “While it is known that natural sources are of paramount importance summarizing 60% of the currently used anticancer agents (Samarghandian et al., 2010), and the in vitro results are promising, in vivo studies are necessary to ultimately confirm a biological effect.” – lines 372-375. Moreover, this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material (most of the collected acorn material was consumed in preliminary / prefeasibility studies for roasting trials, product development, sensory analysis, etc.) and the current work aimed to develop the acorn powder-based product, the in vitro antioxidant and biological properties of acorns being studied. Furthermore, as the reviewer mentioned, for the future work we intend to perform in vivo studies as well, interdisciplinary research with a research group from University of Medicine being sought.

Comment: Line 64: There are a list of countries and it is included Sardinia which is no a country.

Authors response: corrected

Comment: Line 65. A dot should be written between America and Oak. 

Authors response: corrected

Comment: Line 66 cake is redundant since it is mentioned in the line before in a lis of uses of acorns.

Authors response: corrected

Comment: Line 81-82, It is said the there is an antibacterial effect against some bacteria species but a fungal species is included, Candida albicans. It should be expressed in a different way if the acorns also have the same effect against the fungus.

Authors response: corrected

Comment: Statistical Analysis 

Authors said all the analysis were done in duplicate, and they should have been done at least in triplicate to have reliable results. Why didn't you do a triplicate at least?

Authors response: Dear reviewer, the analysis in this research were done in duplicate due to the lack of materials. As previously mentioned, this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material, most of the collected acorn material being consumed in preliminary / prefeasibility studies for roasting trials, product development, sensory analysis, etc.

Comment: The main objetive seem to be the potential use of acorn powder as a substitute of cocoa powder and the results seem to conclude that it is possible based on hedonic test, but there are some test mixed like antibacterial activity, the polyphenols content or the anti proliferative properties that are not really related to the potential use of acorn powder since they are tested in vitro and it is not said what happen in vivo in addition to the fact that all the analysis were done in duplicate.

Authors response: This study was aimed at investigating the acorn derived powder as an alternative for cocoa powder in order to obtain a chocolate that does not contain stimulants of the nervous system. Based on the hedonic test, we showed the degree of acceptability of the new acorn powder based product obtained. Besides the sensory properties of the new confectionery product obtained, it was aimed to perform a preliminary study of the biological and antioxidant properties of the proposed key ingredient – roasted acorn powder in order to show its potential advantages as a future food ingredient; it does not contain stimuli of the nervous system / successfully replacing cocoa and carob powder, has a high content of polyphenols, has the ability to inhibit the growth of Gram-positive bacterium (Bacillus cereus) and have the ability to reduce the cell viability of the HFL-1 (human fetal lung fibroblast) and DLD-1 (colorectal adenocarcinoma) lines.

Comment: Antibacterial properties seem to be only for one bacteria species, which is not enough to affirm that the extract has antibacterial effect.

Authors response: The research studied the antimicrobial effects of aqueous extracts for Salmonella Enteritidis, Salmonella Typhimurium, Staphylococcus aureus, Bacillus cereus. Following the analyzes, it was concluded that aqueous acorn extracts have the ability to inhibit growth of Bacillus cereus. Throughout the manuscript it is specified that it has a limited antimicrobial effect, that is specific for Bacillus cereus.

Comment: Is there any "in vivo" studies to know if the "in vitro" properties work as well in vivo?, since the acorn powder has to be eaten and pass through the digestive system.

Authors response: We agree the reviewer recommendation about the necessity of studying the in vivo, besides the in vitro properties, so for the future work we intend to perform in vivo studies as well, interdisciplinary research with a research group from University of Medicine being sought. As mentioned previously this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material and the current work aimed to develop the acorn powder-based product, in addition, the in vitro antioxidant and biological properties of acorn powder as a key raw material were also studied. We did not find any  studies for extrapolating the in vitro studies. However, there is a pattern of new food products development and characterization studies / published papers, where the techno-functional properties of some acorn based food products was performed: acorn bread [Kim&Joo, 2019; Shishehbor et al., 2020], muffins with added acorn jelly powder and acorn ethanol extract powder [Kim et al., 2012], gluten – free biscuits made from corn – acorn flour [Korus et al., 2017], biscuits with acorn flour [Pasqualone et al., 2019; Amina& Djamel, 2018; Šálková et al., 2011; Parsaei et al., 2018]

Kim, J.-M., & Joo, J-I. (2019). Quality characteristics of acorn bread added with milk. Journal of the Korean Society of Food Culture, 34(3), 343–352. https://doi.org/10.7318/KJFC/2019.34.3.343

Kim, S. H., Lee,W. K., Choi, C. S., & Cho, S. M. (2012). Quality characteristics of muffins with added acorn jelly powder and acorn ethanol extract powder. Journal of the Korean Society of Food Science and Nutrition, 41(3), 369–375.

Korus, A., Gumul, D., Krystyjan, M., Juszczak, L., & Korus, J. (2017). Evaluation of the quality, nutritional value and antioxidant activity of gluten – free biscuits made from corn – acorn flour or corn – hemp flour biscuits. European Food Research and Technology, 243(8), 1429–1438. https://doi.org/10.1007/.s00217-017-2853-y

Pasqualone, A., Makhlouf, F. Z., Barkat, M., Difonzo, G., Summo, C., Squeo, G., & Caponio, F. (2019). Effect of acorn flour on the physico-chemical and sensory properties of biscuits. Heliyon, 5(8), e02242. https://doi.org/10.1016/j.heliyon.2019.e02242

Amina, M., & Djamel, F. (2018). Effect of fermentation and germination treatments on physicochemical and sensory properties of enriched biscuits with acorn flour. Annals. Food Science and Technology, 19(4), 667–674.

Šálková, T., Divišová, M., Kadochová, Š., Delawská, K., Kadlčková, E., Němečková, L., ... Žemličková, A. (2011). Acorns as a food resource. An experiment with acorn preparation. Interdisciplinaria Archaelogica Natural Sciences in Archeology, 2(2), 133–141. https://doi.org/10.24916/iansa.2011.2.6

Shishehbor, F., Salimi, Z., Veissi, M., Malehi, A. S., Shiri-Nasab, M., &Helli, B. (2020). Effect of oak flour on glycemic index and satiety index of white bread. Iranian Red Crescent Medical Journal, 22(1), 1–7. https://doi.org/10.5812/ircmj.95552

Parsaei,M., Goli,M., & Abbasi, H. (2018). Oak flour as a replacement of wheat and corn flour to improve biscuit antioxidant activity. Food Science & Nutrition, 6(2), 253–258. https://doi.org/10.1002/ fsn3.524

 

Reviewer 2

Comment: The article "Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties" presented by Romina Alina (Vlaic) Marc and co-authors is interesting and useful to the community studying food chemistry and nutrients.  In my opinion, this article should be accepted after minor revision.

Authors started with a good introduction to their research with sufficient literature research.

Their methods are well described. They succeed to present their results clearly.

However, there are some minor issues that require attention:

Authors response: Thank you for all encouraging comments.

Comment: Abstract, Page 1, line 29:  authors may delete “in conclusion” if they wish, and start the sentence with “This study suggested…”

Authors response: corrected

Comment: Page 1, line 39:  authors may introduce what LDL stands for.

Authors response: corrected

Comment: Page 3, line 127: “A Na2CO 7.5%”, should be replaced with “Na2CO3”. (sodium bicarbonates formula should be written correctly. It is better to remove “A” at the beginning of the sentence.

Authors response: corrected

Comment: Page 6, in line 234, and in other lines and tables in the articles: authors did not write “°C” in a correct way. Sometimes they did not introduce any spaces “X°C”, sometimes they introduce a space between the value and °C, sometimes they add a space between ° and C, and sometimes they add 2 spaces. Please notice that °C are connected. Authors can write X°C (without space) or X °C (with a space) but should be homogenous within the text.

Authors response: corrected

Comment: Page 8, line 296: rations should be written in a similar way. Either “X:Y” or “X : Y”.

Authors response: corrected

 

Reviewer 3

Manuscript entitled " Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties " presented to evaluate its sensory and nutritional properties as a potential alternative for cocoa powder in the manufacture of chocolate.

Authors response: Thank you for all encouraging comments.

Comment: Introduction section

In this paragraph it is necessary that the authors explain the advantage and disadvantage of optimization of roasting conditions with different time and temperature intervals. Please add references.

Please add and explain the effect of roasting time and roasting temperature on the color of cocoa powder and acorn powder for sensory properties. These effects are pivotal to optimize the roasting operations of cocoa and acorn. Please add references.

Materials and Methods section

Please clearly elucidate the operation weight of the sample when the roasting performance of the acorn is evaluated.

 

Authors response: the selection of parameters (temperature & time) for acorns’ roasting operation was done in an empirical way, while no other similar studies were reported in literature for acorn or other forest seeds. Moreover, the present study might be considered a PIONEERING STUDY for acorn roasting aiming to replace cocoa powder in confectionery products. Consequently, as presented in Table 1, empirically selected temperatures of 135 and 160 °C were studied, the organoleptic profile (color / flavor) being monitored for 90 minutes, samples being analyzed in 10 / 15 min intervals. Based on the same rationale, the higher temperatures (180, 200 and 220 °C) were also selected empirically; however, the sampling time intervals was decreased to 5 min (table 2 and 3) and the total roasting time to 25 min (after this time, material burning was detected). Moreover, the Results and Discussion section 3.1 Optimization of roasting for acorn powder production, presents the obtained results (clear & organized tables with roasting time & temperature parameters / organoleptic profiles) and critically discuss them.

Comment: L 97-98 Please explain deeply the operation of conditioning operation.

Authors response: corrected

Comment: L 99 Please clearly elucidate the condition of humidity and room temperature when roasting procedure was performed.

Authors response: corrected.

Comment: Please add the quantitative measures were performed by means of the International Commission on Illumination (CIE) L*a*b* when the roasting performance of acorn is evaluated.

Results and Discussion

Please clearly elucidate the values of L*, a*, b*, and the color difference ΔE when the roasting performance of acorn is evaluated, before and after roasting operation at 220 oC/15 min.

Please compare the values of L*, a*, b*, and the total color difference ΔE when roasting operation of the acorn at 220 °C/15min with commercial cocoa powder.

Authors response: Dear reviewer, in the experimental design of this research we did not include the instrumental color analysis due to the lack of the available equipment. However, the color was assessed during the organoleptic analysis, the results being presented in Table 1, 2 and 3. Thank you for this suggestion, in the future research we will include this analysis in the experimental design, a colorimeter being recently bought in our laboratory; however, even now the equipment is available in our laboratory, we are not able to perform the instrumental color analysis, while all collected acorn material was consumed.

Thank you once again for your valuable comments. The authors are available if there are any further queries.

--

 

Best regards,

Marc (Vlaic) Romina Alina et al.

Author Response File: Author Response.docx

Reviewer 3 Report

Comments and Suggestions for Authors

     Manuscript entitled " Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties " presented to evaluate its sensory and nutritional properties as a potential alternative for cocoa powder in the manufacture of chocolate.

Introduction section

1. In this paragraph it is necessary that the authors explain the advantage and disadvantage of optimization of roasting conditions with different time and temperature intervals. Please add references.
2. Please add and explain the effect of roasting time and roasting temperature on the color of cocoa powder and acorn powder for sensory properties. These effects are pivotal to optimize the roasting operations of cocoa and acorn. Please add references.

Materials and Methods section

1. Please clearly elucidate the operation weight of the sample when the roasting performance of the acorn is evaluated.
2. L 97-98 Please explain deeply the operation of conditioning operation.
3. L 99 Please clearly elucidate the condition of humidity and room temperature when roasting procedure was performed.
4. Please add the quantitative measures were performed by means of the International Commission on Illumination (CIE) L*a*b* when the roasting performance of acorn is evaluated.

Results and Discussion

1. Please clearly elucidate the values of L*, a*, b*, and the color difference ΔE when the roasting performance of acorn is evaluated, before and after roasting operation at 220 ^oC/15 min.
2. Please compare the values of L*, a*, b*, and the total color difference ΔE when roasting operation of the acorn at 220 ^oC/15min with commercial cocoa powder.

Date of this review

1 AUG 2021

Comments for author File: Comments.pdf

Author Response

Subject: Submission of revised manuscript No. Forests: 1299438

 

Dear Special Issue Editor, Dr. Apostolos P. Kyriazopoulos

Dear Reviewers,

 

Thank you for considering our manuscript, “Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties” submitted for publication in your esteemed Journal Forests MDPI, Special Issue "Non-wood Forest Products".

As suggested by your editorial office, the authors have responded point by point to all the concerns raised by the reviewers. Also enclosed is a detailed description of the revisions made to the manuscript. We have highlighted in the text the changes we made.

We thank the reviewers for their constructive suggestions. In response to their comments, our manuscript has been improved.

Editorial comments:

Reviewer 1

Thank you for all your encouraging comments.

Comment: Introduction:

It should be better explained if the antiproliferative effect which is said to exist in vitro could exist also in vivo, and the same for the antibacterial effect since the author is talking about a product to be eaten. 

Authors response: To the best of our knowledge, to date, there is no supporting data to make any inferences in the introduction about any in vivo antiproliferative effect of red oak fruits. We present in the Results and discussion section of the manuscript (L367-371) data on the antiproliferative activities of other Quercus species. However, based on the reviewer recommendation, we introduced the following sentence in the manuscript “While it is known that natural sources are of paramount importance summarizing 60% of the currently used anticancer agents (Samarghandian et al., 2010), and the in vitro results are promising, in vivo studies are necessary to ultimately confirm a biological effect.” – lines 372-375. Moreover, this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material (most of the collected acorn material was consumed in preliminary / prefeasibility studies for roasting trials, product development, sensory analysis, etc.) and the current work aimed to develop the acorn powder-based product, the in vitro antioxidant and biological properties of acorns being studied. Furthermore, as the reviewer mentioned, for the future work we intend to perform in vivo studies as well, interdisciplinary research with a research group from University of Medicine being sought.

Comment: Line 64: There are a list of countries and it is included Sardinia which is no a country.

Authors response: corrected

Comment: Line 65. A dot should be written between America and Oak. 

Authors response: corrected

Comment: Line 66 cake is redundant since it is mentioned in the line before in a lis of uses of acorns.

Authors response: corrected

Comment: Line 81-82, It is said the there is an antibacterial effect against some bacteria species but a fungal species is included, Candida albicans. It should be expressed in a different way if the acorns also have the same effect against the fungus.

Authors response: corrected

Comment: Statistical Analysis 

Authors said all the analysis were done in duplicate, and they should have been done at least in triplicate to have reliable results. Why didn't you do a triplicate at least?

Authors response: Dear reviewer, the analysis in this research were done in duplicate due to the lack of materials. As previously mentioned, this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material, most of the collected acorn material being consumed in preliminary / prefeasibility studies for roasting trials, product development, sensory analysis, etc.

Comment: The main objetive seem to be the potential use of acorn powder as a substitute of cocoa powder and the results seem to conclude that it is possible based on hedonic test, but there are some test mixed like antibacterial activity, the polyphenols content or the anti proliferative properties that are not really related to the potential use of acorn powder since they are tested in vitro and it is not said what happen in vivo in addition to the fact that all the analysis were done in duplicate.

Authors response: This study was aimed at investigating the acorn derived powder as an alternative for cocoa powder in order to obtain a chocolate that does not contain stimulants of the nervous system. Based on the hedonic test, we showed the degree of acceptability of the new acorn powder based product obtained. Besides the sensory properties of the new confectionery product obtained, it was aimed to perform a preliminary study of the biological and antioxidant properties of the proposed key ingredient – roasted acorn powder in order to show its potential advantages as a future food ingredient; it does not contain stimuli of the nervous system / successfully replacing cocoa and carob powder, has a high content of polyphenols, has the ability to inhibit the growth of Gram-positive bacterium (Bacillus cereus) and have the ability to reduce the cell viability of the HFL-1 (human fetal lung fibroblast) and DLD-1 (colorectal adenocarcinoma) lines.

Comment: Antibacterial properties seem to be only for one bacteria species, which is not enough to affirm that the extract has antibacterial effect.

Authors response: The research studied the antimicrobial effects of aqueous extracts for Salmonella Enteritidis, Salmonella Typhimurium, Staphylococcus aureus, Bacillus cereus. Following the analyzes, it was concluded that aqueous acorn extracts have the ability to inhibit growth of Bacillus cereus. Throughout the manuscript it is specified that it has a limited antimicrobial effect, that is specific for Bacillus cereus.

Comment: Is there any "in vivo" studies to know if the "in vitro" properties work as well in vivo?, since the acorn powder has to be eaten and pass through the digestive system.

Authors response: We agree the reviewer recommendation about the necessity of studying the in vivo, besides the in vitro properties, so for the future work we intend to perform in vivo studies as well, interdisciplinary research with a research group from University of Medicine being sought. As mentioned previously this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material and the current work aimed to develop the acorn powder-based product, in addition, the in vitro antioxidant and biological properties of acorn powder as a key raw material were also studied. We did not find any  studies for extrapolating the in vitro studies. However, there is a pattern of new food products development and characterization studies / published papers, where the techno-functional properties of some acorn based food products was performed: acorn bread [Kim&Joo, 2019; Shishehbor et al., 2020], muffins with added acorn jelly powder and acorn ethanol extract powder [Kim et al., 2012], gluten – free biscuits made from corn – acorn flour [Korus et al., 2017], biscuits with acorn flour [Pasqualone et al., 2019; Amina& Djamel, 2018; Šálková et al., 2011; Parsaei et al., 2018]

Kim, J.-M., & Joo, J-I. (2019). Quality characteristics of acorn bread added with milk. Journal of the Korean Society of Food Culture, 34(3), 343–352. https://doi.org/10.7318/KJFC/2019.34.3.343

Kim, S. H., Lee,W. K., Choi, C. S., & Cho, S. M. (2012). Quality characteristics of muffins with added acorn jelly powder and acorn ethanol extract powder. Journal of the Korean Society of Food Science and Nutrition, 41(3), 369–375.

Korus, A., Gumul, D., Krystyjan, M., Juszczak, L., & Korus, J. (2017). Evaluation of the quality, nutritional value and antioxidant activity of gluten – free biscuits made from corn – acorn flour or corn – hemp flour biscuits. European Food Research and Technology, 243(8), 1429–1438. https://doi.org/10.1007/.s00217-017-2853-y

Pasqualone, A., Makhlouf, F. Z., Barkat, M., Difonzo, G., Summo, C., Squeo, G., & Caponio, F. (2019). Effect of acorn flour on the physico-chemical and sensory properties of biscuits. Heliyon, 5(8), e02242. https://doi.org/10.1016/j.heliyon.2019.e02242

Amina, M., & Djamel, F. (2018). Effect of fermentation and germination treatments on physicochemical and sensory properties of enriched biscuits with acorn flour. Annals. Food Science and Technology, 19(4), 667–674.

Šálková, T., Divišová, M., Kadochová, Š., Delawská, K., Kadlčková, E., Němečková, L., ... Žemličková, A. (2011). Acorns as a food resource. An experiment with acorn preparation. Interdisciplinaria Archaelogica Natural Sciences in Archeology, 2(2), 133–141. https://doi.org/10.24916/iansa.2011.2.6

Shishehbor, F., Salimi, Z., Veissi, M., Malehi, A. S., Shiri-Nasab, M., &Helli, B. (2020). Effect of oak flour on glycemic index and satiety index of white bread. Iranian Red Crescent Medical Journal, 22(1), 1–7. https://doi.org/10.5812/ircmj.95552

Parsaei,M., Goli,M., & Abbasi, H. (2018). Oak flour as a replacement of wheat and corn flour to improve biscuit antioxidant activity. Food Science & Nutrition, 6(2), 253–258. https://doi.org/10.1002/ fsn3.524

 

Reviewer 2

Comment: The article "Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties" presented by Romina Alina (Vlaic) Marc and co-authors is interesting and useful to the community studying food chemistry and nutrients.  In my opinion, this article should be accepted after minor revision.

Authors started with a good introduction to their research with sufficient literature research.

Their methods are well described. They succeed to present their results clearly.

However, there are some minor issues that require attention:

Authors response: Thank you for all encouraging comments.

Comment: Abstract, Page 1, line 29:  authors may delete “in conclusion” if they wish, and start the sentence with “This study suggested…”

Authors response: corrected

Comment: Page 1, line 39:  authors may introduce what LDL stands for.

Authors response: corrected

Comment: Page 3, line 127: “A Na2CO 7.5%”, should be replaced with “Na2CO3”. (sodium bicarbonates formula should be written correctly. It is better to remove “A” at the beginning of the sentence.

Authors response: corrected

Comment: Page 6, in line 234, and in other lines and tables in the articles: authors did not write “°C” in a correct way. Sometimes they did not introduce any spaces “X°C”, sometimes they introduce a space between the value and °C, sometimes they add a space between ° and C, and sometimes they add 2 spaces. Please notice that °C are connected. Authors can write X°C (without space) or X °C (with a space) but should be homogenous within the text.

Authors response: corrected

Comment: Page 8, line 296: rations should be written in a similar way. Either “X:Y” or “X : Y”.

Authors response: corrected

 

Reviewer 3

Manuscript entitled " Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties " presented to evaluate its sensory and nutritional properties as a potential alternative for cocoa powder in the manufacture of chocolate.

Authors response: Thank you for all encouraging comments.

Comment: Introduction section

In this paragraph it is necessary that the authors explain the advantage and disadvantage of optimization of roasting conditions with different time and temperature intervals. Please add references.

Please add and explain the effect of roasting time and roasting temperature on the color of cocoa powder and acorn powder for sensory properties. These effects are pivotal to optimize the roasting operations of cocoa and acorn. Please add references.

Materials and Methods section

Please clearly elucidate the operation weight of the sample when the roasting performance of the acorn is evaluated.

 

Authors response: the selection of parameters (temperature & time) for acorns’ roasting operation was done in an empirical way, while no other similar studies were reported in literature for acorn or other forest seeds. Moreover, the present study might be considered a PIONEERING STUDY for acorn roasting aiming to replace cocoa powder in confectionery products. Consequently, as presented in Table 1, empirically selected temperatures of 135 and 160 °C were studied, the organoleptic profile (color / flavor) being monitored for 90 minutes, samples being analyzed in 10 / 15 min intervals. Based on the same rationale, the higher temperatures (180, 200 and 220 °C) were also selected empirically; however, the sampling time intervals was decreased to 5 min (table 2 and 3) and the total roasting time to 25 min (after this time, material burning was detected). Moreover, the Results and Discussion section 3.1 Optimization of roasting for acorn powder production, presents the obtained results (clear & organized tables with roasting time & temperature parameters / organoleptic profiles) and critically discuss them.

Comment: L 97-98 Please explain deeply the operation of conditioning operation.

Authors response: corrected

Comment: L 99 Please clearly elucidate the condition of humidity and room temperature when roasting procedure was performed.

Authors response: corrected.

Comment: Please add the quantitative measures were performed by means of the International Commission on Illumination (CIE) L*a*b* when the roasting performance of acorn is evaluated.

Results and Discussion

Please clearly elucidate the values of L*, a*, b*, and the color difference ΔE when the roasting performance of acorn is evaluated, before and after roasting operation at 220 oC/15 min.

Please compare the values of L*, a*, b*, and the total color difference ΔE when roasting operation of the acorn at 220 °C/15min with commercial cocoa powder.

Authors response: Dear reviewer, in the experimental design of this research we did not include the instrumental color analysis due to the lack of the available equipment. However, the color was assessed during the organoleptic analysis, the results being presented in Table 1, 2 and 3. Thank you for this suggestion, in the future research we will include this analysis in the experimental design, a colorimeter being recently bought in our laboratory; however, even now the equipment is available in our laboratory, we are not able to perform the instrumental color analysis, while all collected acorn material was consumed.

Thank you once again for your valuable comments. The authors are available if there are any further queries.

--

 

Best regards,

Marc (Vlaic) Romina Alina et al.

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

The problem of this work is, in my opinion, the performance in duplicate of the assays. Although the authors said they did not have enough material for performing the assays in triplicate, it is a big concern since statistically it is not possible to demonstrate anything. The authors should have kept material to properly perform the assays. 

Author Response

Subject: Submission of revised manuscript No. Forests: 1299438

 

Dear Special Issue Editor, Dr. Apostolos P. Kyriazopoulos,

Dear Reviewer,

Thank you for considering our manuscript, “Red oak (Quercus rubra L.) fruits as potential alternative for cocoa powder: optimization of roasting conditions, antioxidant, and biological properties” submitted for publication in the journal Forests MDPI, Special Issue "Non-wood Forest Products".

As suggested by your editorial office, the authors have responded to the last concern raised by the reviewer. We have highlighted in the text the changes we made.

Reviewer 1

Comment: The problem of this work is, in my opinion, the performance in duplicate of the assays. Although the authors said they did not have enough material for performing the assays in triplicate, it is a big concern since statistically it is not possible to demonstrate anything. The authors should have kept material to properly perform the assays.

Authors response: Dear reviewer, the analysis in this research were done in duplicate (not triplicate) REPLICATIONS (different aqueous extractions) due to the lack of materials, as mentioned in our previous response. For each sample, 2 aqueous extractions (2 REPLICATIONS of experiment) were performed and, each one, was measured in 2 REPETITIONS (2 replications x 2 repetitions, a total of 4 measurements for each sample). In this view, we consider that a duplicate replications experiment with 2 repetitions measurements for each replication, is sufficient to statistically demonstrate the differences (if any) among analyzed samples. As previously mentioned, this is the first study of our group on red oak acorn valorization and potential use as a food ingredient; this initial research had a limited available material, most of the collected acorn material being consumed in preliminary / prefeasibility studies for roasting trials, product development, sensory analysis, etc. Moreover, the present study might be considered a PIONEERING STUDY for acorn roasting aiming to replace cocoa powder in confectionery products, to the best of our knowledge, to date, there are no similar studies.

Thank you for pointing out this issue – for making the text clearer, we added on the Section Materials and Methods, Sub-section 2.6. Statistical Analysis, line 202 the term “replications” besides duplicate.

Best regards,

Marc (Vlaic) Romina Alina et al.

Author Response File: Author Response.docx