Functional Properties of High-Pressure Assisted Enzymatic Tamarind Kernel Protein Hydrolysate and Foam-Mat Powder Characteristics as Affected by HPMC Concentration and Drying Temperature

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript addresses a relevant and timely topic in food science by investigating the valorization of TKP. The systematic evaluation of functional properties and the optimization of foam‑mat drying conditions are strengths of the study. In addition, the comparative assessment against EW and SPI provides useful context for interpreting the functional performance of TKP‑based materials. Overall, the manuscript is generally well written, however, a closer examination reveals several issues that should be addressed to strengthen its scientific rigor and practical relevance.

1. The current title implies a comprehensive optimization study, however, the limited evaluation of only two HPMC concentrations and three drying temperatures reflects preliminary screening rather than rigorous optimization. To avoid overstating the scope, the title should be revised to indicate an exploratory or preliminary investigation.
2. The manuscript doesn't clearly describe the basis for sample size selection, nor does it report any statistical power analysis. Furthermore, the rationale for choosing specific experimental parameter ranges, such as HPMC concentrations (1% and 1.5%) and the selected foam‑mat drying temperatures, remains unclear. Providing justification for these choices, whether based on preliminary trials, literature precedent, or process constraints, would enhance the transparency and robustness of the study.
3. Although the authors report notable changes in functional properties following HPP and enzymatic hydrolysis, the discussion lacks sufficient mechanistic depth. Greater insight into molecular‑level phenomena, such as protein unfolding, aggregation, or peptide formation, is needed to explain the observed functional outcomes. Strengthening the link between structural modifications and functional behavior, ideally supported by additional characterization techniques (e.g., secondary structure analysis, molecular weight distribution), would substantially improve the scientific contribution.
4. The manuscript doesn't adequately address the practical implications of applying HPP‑assisted hydrolysis and foam‑mat drying at an industrial scale. Key considerations such as cost‑benefit analysis, comparative energy consumption, processing time, and potential scale‑up challenges should be discussed.
5. Important quality parameters including bulk density, particle size distribution, reconstitution behavior, hygroscopicity, and storage stability are not evaluated. These attributes are essential for assessing the usability and commercial viability of the dried powder in real food applications and should be considered in future revisions.
6. Comparison of functional properties is limited to EW and SPI. Including comparisons with commercially available protein hydrolysates or other plant‑based protein alternatives would provide a more meaningful benchmark for evaluating the novelty, competitiveness, and application potential of TKP‑derived hydrolysates.
7. Several methodological aspects require further clarification, including the degree of hydrolysis achieved, the peptide molecular weight distribution, and the reproducibility of HPP treatment conditions. Moreover, the foam‑mat drying optimization appears limited, as only three drying temperatures were examined. Exploration of additional critical variables, such as foam thickness, air velocity, and drying air humidity, would yield a more comprehensive optimization framework.
8. The manuscript would benefit from a clearer discussion of the practical implications of the reported functional property enhancements. Establishing threshold values or performance benchmarks that justify industrial adoption would help translate the findings from laboratory‑scale observations to application‑oriented conclusions.

Author Response

Reviewer 1:

Comments and Suggestions for Authors

The manuscript addresses a relevant and timely topic in food science by investigating the valorization of TKP. The systematic evaluation of functional properties and the optimization of foam‑mat drying conditions are strengths of the study. In addition, the comparative assessment against EW and SPI provides useful context for interpreting the functional performance of TKP‑based materials. Overall, the manuscript is generally well written, however, a closer examination reveals several issues that should be addressed to strengthen its scientific rigor and practical relevance.

Response: We sincerely thank the reviewer for the constructive comments and valuable suggestions. We have carefully considered the suggested improvements and revised the manuscript accordingly, as highlight in the yellow color.

 

1. The current title implies a comprehensive optimization study, however, the limited evaluation of only two HPMC concentrations and three drying temperatures reflects preliminary screening rather than rigorous optimization. To avoid overstating the scope, the title should be revised to indicate an exploratory or preliminary investigation.

Response: Thank you for your suggestion, we revised in the conclusion line 701 and Line 703-710.

 

2. The manuscript doesn't clearly describe the basis for sample size selection, nor does it report any statistical power analysis. Furthermore, the rationale for choosing specific experimental parameter ranges, such as HPMC concentrations (1% and 1.5%) and the selected foam‑mat drying temperatures, remains unclear. Providing justification for these choices, whether based on preliminary trials, literature precedent, or process constraints, would enhance the transparency and robustness of the study.

Response: Based on preliminary observations, lower HPMC concentrations resulted in unstable foams, while higher concentrations caused excessive viscosity that hindered effective foaming and spreading. Accordingly, the drying temperatures were selected to reflect moderate foam-mat drying conditions that help avoid excessive thermal degradation.

 

3. Although the authors report notable changes in functional properties following HPP and enzymatic hydrolysis, the discussion lacks sufficient mechanistic depth. Greater insight into molecular‑level phenomena, such as protein unfolding, aggregation, or peptide formation, is needed to explain the observed functional outcomes. Strengthening the link between structural modifications and functional behavior, ideally supported by additional characterization techniques (e.g., secondary structure analysis, molecular weight distribution), would substantially improve the scientific contribution.

Response: We sincerely thank the reviewer for the thoughtful and constructive comments. Although mechanistic aspects underlying the changes in functional properties following HPP and enzymatic hydrolysis were discussed in several sections of the discussion, for example in Line 378-379, 428-431. We greatly appreciate the reviewer’s valuable suggestion to further emphasize the link between structural modifications and functional behavior. This insightful recommendation will be highly beneficial in guiding our future studies and in enabling a deeper mechanistic investigation in our subsequent work.

 

4. The manuscript doesn't adequately address the practical implications of applying HPP‑assisted hydrolysis and foam‑mat drying at an industrial scale. Key considerations such as cost‑benefit analysis, comparative energy consumption, processing time, and potential scale‑up challenges should be discussed.

Response: We sincerely appreciate the reviewer’s important comment and agree that further discussion of industrial-scale implications would enhance the manuscript. Accordingly, the conclusion has been expanded to highlight key practical considerations, including qualitative cost–benefit aspects, energy use, processing time, and scale-up challenges related to HPP-assisted hydrolysis and foam-mat drying.

 

5. Important quality parameters including bulk density, particle size distribution, reconstitution behavior, hygroscopicity, and storage stability are not evaluated. These attributes are essential for assessing the usability and commercial viability of the dried powder in real food applications and should be considered in future revisions.

Response: As the present study was designed as an exploratory investigation focusing on functional enhancement and foam-mat drying feasibility, these quality parameters were beyond its current scope. This limitation has now been acknowledged in the revised manuscript, and the suggested analyses are identified as important directions for future research aimed at product development and practical application.

 

6. Comparison of functional properties is limited to EW and SPI. Including comparisons with commercially available protein hydrolysates or other plant‑based protein alternatives would provide a more meaningful benchmark for evaluating the novelty, competitiveness, and application potential of TKP‑derived hydrolysates.

Response: Egg white (EW) and soy protein isolate (SPI) were selected as representative animal- and plant-derived proteins, respectively, owing to their excellent functional properties.

 

7. Several methodological aspects require further clarification, including the degree of hydrolysis achieved, the peptide molecular weight distribution, and the reproducibility of HPP treatment conditions. Moreover, the foam‑mat drying optimization appears limited, as only three drying temperatures were examined. Exploration of additional critical variables, such as foam thickness, air velocity, and drying air humidity, would yield a more comprehensive optimization framework.

Response: We added the degree of hydrolysis in the revised manuscript Line 168. The HPP treatments were carried out under consistent processing conditions to ensure reproducibility. For foam-mat drying, we acknowledge that the optimization was limited to drying temperature and therefore represents an initial screening rather than a full optimization. Additional parameters such as foam thickness, air velocity, and drying air humidity are important considerations and are now highlighted as key areas for future study.

 

8. The manuscript would benefit from a clearer discussion of the practical implications of the reported functional property enhancements. Establishing threshold values or performance benchmarks that justify industrial adoption would help translate the findings from laboratory‑scale observations to application‑oriented conclusions.

Response: As response in 4.

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Authors, you should address the comments highlighted across the text, tables and figures.

Comments for author File: Comments.pdf

Author Response

We sincerely thank the reviewer for the comments and suggestions. We have carefully considered the suggested improvements and revised the manuscript accordingly as highlight in the green color.

Reviewer 3 Report

Comments and Suggestions for Authors

This manuscript addresses an important and timely topic in food science by valorizing tamarind kernel powder, an underutilized agro-industrial by-product, through high-pressure processing assisted enzymatic hydrolysis and foam-mat drying. The study is well designed and integrates protein chemistry, process engineering, and functional food evaluation. A major strength lies in the systematic comparison of TKP-HD with well-established reference proteins, egg white and soy protein isolate, which allows readers to directly assess industrial relevance. The combined evaluation of functional properties, antioxidant activity, and microstructure provides a comprehensive understanding of how processing conditions influence performance. The manuscript is generally well written, methodologically sound, and supported by an extensive and relevant literature base. With these merits in mind, I provide several suggestions below to further strengthen the manuscript.

1. Clarification of novelty relative to prior work
Although novelty is stated in the Introduction (Lines 102–105), the distinction from previous HPP studies on tamarind proteins remains subtle. I suggest explicitly clarifying, in the Introduction and Conclusions, how the combined evaluation of functional benchmarking and foam-mat drying advances beyond Ref. [4; ‘The effects of high pressure-assisted enzymatic hydrolysis on degree of hydrolysis and antioxidant properties of tamarind kernel protein’].

2. Statistical reporting and experimental design
Section 2.8 states that one-way ANOVA was used, which is acceptable. However, several results (Figures 3 and 4, Table 4) involve two experimental factors varied simultaneously, namely drying temperature and HPMC concentration. Please clarify how these factors were handled statistically. Specifically, indicate whether interaction effects were evaluated, or whether each temperature–HPMC combination was treated as an independent group in a one-way ANOVA. If interaction effects were not assessed, please state this explicitly in Section 2.8 to avoid ambiguity.

3. Protein content normalization in functional comparisons
Functional properties of TKP-HD are compared with EW and SPI despite large differences in protein content (Table 2). Please discuss this limitation more explicitly and consider adding a brief justification that results reflect ingredient-level functionality rather than protein-normalized performance.

4. Mechanistic interpretation of antioxidant activity
The discussion attributes antioxidant activity primarily to peptides and Maillard reaction products (Section 3.2.2). I recommend tempering causal language and rephrasing to indicate associative rather than definitive mechanisms, unless direct peptide profiling is available.

5. In Table 3, briefly indicate how hydrophobic and hydrophilic amino acids were classified.

6. Improve figure readability by increasing font size of axis labels in Figures 1–4.

Overall, this is a solid and meaningful contribution to the field of food protein processing and functional ingredient development. The manuscript would benefit from moderate clarification and refinement, but no additional experiments appear necessary. I therefore recommend Major Revision, and I encourage the authors to address the comments above to further enhance clarity, rigor, and impact. I commend the authors for their comprehensive and well-executed study.

Author Response

Reviewer 3:

This manuscript addresses an important and timely topic in food science by valorizing tamarind kernel powder, an underutilized agro-industrial by-product, through high-pressure processing assisted enzymatic hydrolysis and foam-mat drying. The study is well designed and integrates protein chemistry, process engineering, and functional food evaluation. A major strength lies in the systematic comparison of TKP-HD with well-established reference proteins, egg white and soy protein isolate, which allows readers to directly assess industrial relevance. The combined evaluation of functional properties, antioxidant activity, and microstructure provides a comprehensive understanding of how processing conditions influence performance. The manuscript is generally well written, methodologically sound, and supported by an extensive and relevant literature base. With these merits in mind, I provide several suggestions below to further strengthen the manuscript.

Response: We sincerely thank the reviewer for the constructive comments and valuable suggestions. We have carefully considered the suggested improvements and revised the manuscript accordingly, as highlight in the blue color.

 

1. Clarification of novelty relative to prior work

Although novelty is stated in the Introduction (Lines 102–105), the distinction from previous HPP studies on tamarind proteins remains subtle. I suggest explicitly clarifying, in the Introduction and Conclusions, how the combined evaluation of functional benchmarking and foam-mat drying advances beyond Ref. [4; ‘The effects of high pressure-assisted enzymatic hydrolysis on degree of hydrolysis and antioxidant properties of tamarind kernel protein’].

Response: We thank the reviewer for this valuable comment. While Reference [4] provided a foundational understanding of the hydrolysis degree and antioxidant properties of TKP-HD, our current work aims to bridge the gap between bioactivity and practical application. In addition, we revised in the introduction, as in Line 107-114.

 

2. Statistical reporting and experimental design

Section 2.8 states that one-way ANOVA was used, which is acceptable. However, several results (Figures 3 and 4, Table 4) involve two experimental factors varied simultaneously, namely drying temperature and HPMC concentration. Please clarify how these factors were handled statistically. Specifically, indicate whether interaction effects were evaluated, or whether each temperature–HPMC combination was treated as an independent group in a one-way ANOVA. If interaction effects were not assessed, please state this explicitly in Section 2.8 to avoid ambiguity.

Response: In the present study, all the experiments were performed following a completely randomized design (CRD), as mentioned in 2.8. The experiments involving both drying temperature and HPMC concentration were analyzed by treating each temperature–HPMC combination as an independent group and applying one-way ANOVA followed by Duncan’s multiple range test. Interaction effects between drying temperature and HPMC concentration were not evaluated.

 

3. Protein content normalization in functional comparisons

Functional properties of TKP-HD are compared with EW and SPI despite large differences in protein content (Table 2). Please discuss this limitation more explicitly and consider adding a brief justification that results reflect ingredient-level functionality rather than protein-normalized performance.

Response: We edited in the revised manuscript, Line 174-177.

 

4. Mechanistic interpretation of antioxidant activity

The discussion attributes antioxidant activity primarily to peptides and Maillard reaction products (Section 3.2.2). I recommend tempering causal language and rephrasing to indicate associative rather than definitive mechanisms, unless direct peptide profiling is available.

Response: Thank you for your valuable comment and we revised in Line 555-557.

 

5. In Table 3, briefly indicate how hydrophobic and hydrophilic amino acids were classified.

Response: The hydrophobic and hydrophilic amino acids were classified by the polarity of side chain groups.

 

6. Improve figure readability by increasing font size of axis labels in Figures 1-4.

Response: We edited the Figures 1-4, in the revised manuscript.

 

Overall, this is a solid and meaningful contribution to the field of food protein processing and functional ingredient development. The manuscript would benefit from moderate clarification and refinement, but no additional experiments appear necessary. I therefore recommend Major Revision, and I encourage the authors to address the comments above to further enhance clarity, rigor, and impact. I commend the authors for their comprehensive and well-executed study.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The authors’ responses demonstrate commendable effort; however, several critical concerns remain unresolved. To strengthen the manuscript, speficic and actionable suggestions are outlined below.

1. The current title suggests comprehensive "optimization," yet the study explores only two HPMC concentrations and three drying temperatures. This scope is more consistent with preliminary screening rather than full optimization. To better reflect the nature of the work, the authors consider revising the title so that it clearly conveys its exploratory or preliminary character. This adjustment will help set appropriate expectations for readers and highlight the value of the study's initial findings.
2. The authors acknowledged my concern but did not provide new experimental data. Instead, the response referenced existing discussion, which remains largely descriptive rather than mechanistic. To strengthen the manuscript, the authors should explain mechanistically how HPP unfolds proteins and exposes enzyme cleavage sites. Also, the authors are encouraged to link peptide characteristics (such as size and hydrophobicity) to the functional properties observed (solubility, oil binding, foaming). This will enhance the rigor of the discussion and provide readers with clearer mechanistic insights.
3. The authors compared TKP-HD against native EW and SPI but didn't include commercially available protein hydrolysates, which represent the most direct competitors. To strengthen the manuscript, the authors should either add comparison data for one or two commercial hydrolysates (such as soy, pea, or rice) and evaluate solubility, oil holding capacity, antioxidant activity, and foam capacity to position TKP-HD relative to existing products, or provide a detailed written justification explaining why comparison with EW and SPI alone is scientifically sufficient. The current restatement of the original selection does not adequately address this concern.

Author Response

Reviewer 1

The authors’ responses demonstrate commendable effort; however, several critical concerns remain unresolved. To strengthen the manuscript, specific and actionable suggestions are outlined below.

Response: We sincerely thank the reviewer for the thoughtful and constructive comments.

 

1. The current title suggests comprehensive "optimization," yet the study explores only two HPMC concentrations and three drying temperatures. This scope is more consistent with preliminary screening rather than full optimization. To better reflect the nature of the work, the authors consider revising the title so that it clearly conveys its exploratory or preliminary character. This adjustment will help set appropriate expectations for readers and highlight the value of the study's initial findings.

Response: We revised the title as “Functional Properties of High-pressure Assisted Enzymatic Tamarind Kernel Protein Hydrolysate and Foam-mat Powder Characteristics as Affected by HPMC Concentration and Drying Temperature”

2. The authors acknowledged my concern but did not provide new experimental data. Instead, the response referenced existing discussion, which remains largely descriptive rather than mechanistic. To strengthen the manuscript, the authors should explain mechanistically how HPP unfolds proteins and exposes enzyme cleavage sites. Also, the authors are encouraged to link peptide characteristics (such as size and hydrophobicity) to the functional properties observed (solubility, oil binding, foaming). This will enhance the rigor of the discussion and provide readers with clearer mechanistic insights.

Response: We have revised and explained in the “Introduction”, as in Line 85-97.

3. The authors compared TKP-HD against native EW and SPI but didn't include commercially available protein hydrolysates, which represent the most direct competitors. To strengthen the manuscript, the authors should either add comparison data for one or two commercial hydrolysates (such as soy, pea, or rice) and evaluate solubility, oil holding capacity, antioxidant activity, and foam capacity to position TKP-HD relative to existing products, or provide a detailed written justification explaining why comparison with EW and SPI alone is scientifically sufficient. The current restatement of the original selection does not adequately address this concern.

Response: Scientifically, this study was not intended to position TKP-HD as a direct substitute for commercial protein hydrolysates. Rather, it aimed to determine whether TKP-HD, produced via HPP-assisted enzymatic hydrolysis, can achieve functional performance comparable to that of well-established, high-functionality proteins. In this context, EW and SPI serve as upper-bound functional benchmarks rather than direct market competitors.

          We revised as in Line 184-188, “The functional properties of TKP-HD were compared, on an ingredient-level basis, with egg white (EW) and soy protein isolate (SPI), which are widely used reference proteins representing animal- and plant-derived protein sources. EW and SPI were used as upper-bound functional benchmarks for ingredient-level evaluation of TKP-HD”, in the revised manuscript.

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Authors, the manuscript has been sufficiently improved and, therefore, it can be accepted for publication in Foods in present form, in my opinion.

Author Response

We sincerely thank the reviewer for the time and effort devoted to evaluating our manuscript and for the constructive comments, which have greatly improved the quality of our work.

Reviewer 3 Report

Comments and Suggestions for Authors

Dear Authors,
Thank you for your efforts to revise and improve the manuscript. All issues have been carefully addressed, and I have no further comments or suggestions.

Author Response

We sincerely thank the reviewer for the time and effort devoted to evaluating our manuscript and for the constructive comments, which have greatly improved the quality of our work.