Development and Characterization of a Plant-Based Chicken Nugget Analogue Based on Extruded Sacha Inchi Cake, Textured Soy Protein, and Wheat Gluten

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

REVIEWER COMMENTS AND SUGGESTIONS FOR AUTHORS
Manuscript: "Development and Characterization of a Plant-Based Chicken Nugget Analogue Based on Extruded Sacha Inchi Cake, Textured Soy Protein, and Wheat Gluten"

Dear Authors,

Thank you for submitting your manuscript. The study addresses a relevant and timely topic and the experimental framework is well conceived. The novel incorporation of extruded Sacha Inchi cake (MASI) into a plant-based nugget system is a genuine contribution to the literature, and the comprehensive sensory methodology (CATA, hedonic test, penalty analysis) is commendable. However, the following issues must be addressed before the manuscript can be considered for publication. Please provide a point-by-point response to each numbered comment in your revision letter.

CRITICAL COMMENTS — Must be corrected before re-review

Comment 1 | Abstract / Table 9 | CRITICAL
The carbohydrate content of the optimal formulation is reported as 35.54% in the abstract but as 25.54% in Table 9. These values are mutually inconsistent.
Requested action: Identify and correct the erroneous value. Ensure the correct value is used consistently in the abstract, Table 9, and all related discussion text.

Comment 2 | All experimental sections | CRITICAL
TPA measurements and proximate analyses are reported with n = 2 replicates. This is below the minimum generally required in food science research (n ≥ 3 per AOAC guidelines) and limits the statistical power of all comparisons drawn in the manuscript.
Requested action: Either increase the number of replicates to n ≥ 3 and update all affected tables and statistical outputs, or provide a rigorous statistical justification for the adequacy of n = 2 in this specific experimental context.

MAJOR COMMENTS — Must be addressed in the revision

Comment 3 | Tables 3 and 6 — Statistical model quality | MAJOR
Several fitted models exhibit low explanatory power (cohesiveness: R² = 0.30; cooking loss/yield: R² = 0.36) and statistically significant lack-of-fit (hardness: p = 0.028; cohesiveness: p < 0.0001; chewiness: p = 0.0003). These statistics indicate that the linear mixture model is an inadequate description of the true response surface for these variables. The current discussion minimizes these limitations.
Requested action: Expand the discussion to explicitly acknowledge these limitations and discuss which uncontrolled factors may be contributing to unexplained variance.

Comment 4 | Table 10 — Water activity implications | MAJOR
The water activity of the optimal formulation (aw = 0.986) is significantly higher than that of the commercial reference SF (aw = 0.956), and the difference is statistically significant. This aw value falls within the range that supports the growth of most spoilage microorganisms and several foodborne pathogens (e.g., Salmonella spp. minimum aw ~0.94; Listeria monocytogenes minimum aw ~0.92). The microbiological safety and shelf-life implications of this finding are not discussed in the manuscript.
Requested action: Include a dedicated paragraph discussing the microbiological stability implications of the high aw value and potential mitigation strategies (e.g., freezing, modified atmosphere packaging, antimicrobial hurdles).

MINOR COMMENTS

Comment 5 | Abstract
The number of panelists is not stated in the abstract.
Requested action: Add the number of panelists (n = 70) to the abstract.

Comment 6 | References
References [95] and [96] appear to cite the same work (Taipe et al., 2022, Ciencia Tecnología Agropecuaria 23). This duplicate reduces the credibility of the literature review.
Requested action: Remove the duplicate reference and renumber all subsequent citations accordingly.

Author Response

REVIEWER 1

REVIEWER COMMENTS AND SUGGESTIONS FOR AUTHORS

Manuscript: "Development and Characterization of a Plant-Based Chicken Nugget Analogue Based on Extruded Sacha Inchi Cake, Textured Soy Protein, and Wheat Gluten"
Dear Authors, Thank you for submitting your manuscript. The study addresses a relevant and timely topic and the experimental framework is well conceived. The novel incorporation of extruded Sacha Inchi cake (MASI) into a plant-based nugget system is a genuine contribution to the literature, and the comprehensive sensory methodology (CATA, hedonic test, penalty analysis) is commendable. However, the following issues must be addressed before the manuscript can be considered for publication. Please provide a point-by-point response to each numbered comment in your revision letter.

First of all, thank you very much for all your appreciation of our manuscript by the reviewer. In “blue font”, you will find our answers to your major suggestions/queries about the research.

CRITICAL COMMENTS — Must be corrected before re-review

Comment 1 | Abstract / Table 9 | CRITICAL

The carbohydrate content of the optimal formulation is reported as 35.54% in the abstract but as 25.54% in Table 9. These values are mutually inconsistent.
Requested action: Identify and correct the erroneous value. Ensure the correct value is used consistently in the abstract, Table 9, and all related discussion text.

RESPONSE 1: We thank the reviewer for identifying this inconsistency. After revising the manuscript, it was confirmed that the correct carbohydrate content value for the optimal formulation is 25.54%, as reported in Table 9 (line 551). The value indicated in the abstract (35.54%) corresponded to a transcription error.

In response to this observation:

• The value in the abstract (line 27) was corrected, replacing 35.54% with 25.54%.
• The consistency of this value was verified throughout the manuscript, including the corresponding results and discussion sections.

This discrepancy does not affect the experimental results or the conclusions of the study, since all analyses were conducted using the correct value.

Comment 2 | All experimental sections | CRITICAL

TPA measurements and proximate analyses are reported with n = 2 replicates. This is below the minimum generally required in food science research (n ≥ 3 per AOAC guidelines) and limits the statistical power of all comparisons drawn in the manuscript.
Requested action: Either increase the number of replicates to n ≥ 3 and update all affected tables and statistical outputs, or provide a rigorous statistical justification for the adequacy of n = 2 in this specific experimental context.

RESPONSE 2: We thank the reviewer for this critical observation regarding the number of experimental replicates. In response to this comment, a detailed clarification was incorporated into the “Statistical Analysis” section (Section 3.7, lines 979–991), where it is specified that each nugget formulation evaluated in the study was prepared in two independent batches (batch 1 and batch 2), which were considered experimental units (n = 2). Within each batch, multiple technical replicates (≥3 measurements per parameter) were performed, from which a representative mean value was obtained, with the final value corresponding to the average of both batches.

Additionally, information regarding experimental variability was included, indicating that the coefficients of variation (CV) among replicates were mostly low (generally <15% and in several cases <10%), supporting the reproducibility of the measurements. Nevertheless, it is acknowledged that higher values were observed in some processing-sensitive variables, which is expected in complex food matrices and may be attributed to uncontrolled factors during sample preparation and measurement.

Regarding the proximate analyses of the raw materials used in the study and the optimal nugget formulation, it was specified that these analyses were performed using standardized methods by a specialized external laboratory, with two determinations per sample (n = 2), in accordance with the information provided in the manuscript.

Furthermore, the wording of the footnotes corresponding to the experimental results tables was modified (Table 2, line 248; Table 5, line 399; Table 8, line 531; Table 10, line 592), where it is now explicitly stated that the data correspond to “n = 2 independent batches, with at least three technical replicates per batch,” in order to clarify the experimental structure and avoid ambiguities in data interpretation.

The number of evaluated batches was conditioned by the availability of raw materials during the experimental development.

MAJOR COMMENTS — Must be addressed in the revision

Comment 3 | Tables 3 and 6 — Statistical model quality | MAJOR

Several fitted models exhibit low explanatory power (cohesiveness: R² = 0.30; cooking loss/yield: R² = 0.36) and statistically significant lack-of-fit (hardness: p = 0.028; cohesiveness: p < 0.0001; chewiness: p = 0.0003). These statistics indicate that the linear mixture model is an inadequate description of the true response surface for these variables. The current discussion minimizes these limitations.

Requested action: Expand the discussion to explicitly acknowledge these limitations and discuss which uncontrolled factors may be contributing to unexplained variance.

RESPONSE 3: We thank the reviewer for this valuable observation regarding the quality of fit of the statistical models. In response to this comment, the results and discussion section was revised and expanded (lines 236–262), incorporating a more explicit interpretation of the scope and limitations of the mixture models employed.

First, it is explicitly acknowledged that some variables, such as cohesiveness, cooking loss, cooking yield, and water activity, exhibited low explanatory power (R² ≤ 0.36), which limits their descriptive capability within the mixture space. Consequently, these variables were excluded from the final modeling and from the equations presented (Table 4, line 323), thereby avoiding interpretations based on models with insufficient fit.

Furthermore, it was clarified that the selected models (hardness, springiness, chewiness, and pH) were chosen based on a more consistent behavior within the system, evaluated through a combination of statistical indicators (R², R²adj, R²pred, PRESS, and lack of fit), rather than solely on the coefficient of determination. This methodological decision has been incorporated into the manuscript (lines 264–271).

Additionally, a more extensive discussion regarding the presence of a significant lack of fit in some responses was incorporated, attributing its significance to the limited ability of linear models to capture the complexity of the system. In this regard, it was specified that the use of low-order models was intended to maintain the interpretability of component effects, acknowledging that higher-order models could improve the statistical fit, although at the expense of greater interpretative complexity than what may occur in real phenomena (lines 289–301).

Within this context, the models were reinterpreted as descriptive approximations of system behavior, useful for identifying general trends among mixture components rather than as strict predictive tools.

Subsequently, the discussion was expanded to address potential sources of unexplained variability, including uncontrolled factors such as mixture heterogeneity, ingredient distribution, processing conditions, and instrumental variability, which are inherent to complex food matrices (lines 302–307).

Finally, these clarifications also led to modifications in Table 4 (lines 308–325), which presents the modeling equations for the selected variables (hardness, springiness, chewiness, and pH), as well as revisions to the discussion of Figure 2 corresponding to the Cox trace plots (lines 328–386).

Comment 4 | Table 10 — Water activity implications | MAJOR

The water activity of the optimal formulation (aw = 0.986) is significantly higher than that of the commercial reference SF (aw = 0.956), and the difference is statistically significant. This aw value falls within the range that supports the growth of most spoilage microorganisms and several foodborne pathogens (e.g., Salmonella spp. minimum aw ~0.94; Listeria monocytogenes minimum aw ~0.92). The microbiological safety and shelf-life implications of this finding are not discussed in the manuscript.
Requested action: Include a dedicated paragraph discussing the microbiological stability implications of the high aw value and potential mitigation strategies (e.g., freezing, modified atmosphere packaging, antimicrobial hurdles).

RESPONSE 4: We thank the reviewer for this observation. In response to this observation, a new paragraph was incorporated into the results and discussion section (lines 574–590), explicitly addressing the microbiological stability implications associated with the a_w value (0.986). It is acknowledged that this value falls within the range that favors the growth of spoilage microorganisms and certain foodborne pathogens, representing a potential limitation in terms of shelf life and food safety under uncontrolled conditions.

Furthermore, an explanation was added to contextualize the system under frozen storage conditions (−18 °C), indicating that a significant fraction of the water remains in a solid state, thereby reducing water availability and limiting microbial activity. Nevertheless, it is clarified that this effect is temperature-dependent and reversible after thawing; therefore, microbiological stability cannot be attributed solely to this factor.

Additionally, the discussion was expanded to include complementary preservation strategies, such as continuous frozen storage, modified atmosphere packaging, and the potential use of antimicrobial barriers, in order to improve product safety and extend shelf life.

These additions broaden the discussion beyond the initial formulation-focused approach, providing a more comprehensive interpretation of the product behavior in terms of microbiological stability.

MINOR COMMENTS

Comment 5 | Abstract

The number of panelists is not stated in the abstract.

Requested action: Add the number of panelists (n = 70) to the abstract.

RESPONSE 5: We thank the reviewer for this observation. In response to this observation, the number of panelists was incorporated into the abstract by adding “(n = 70 panelists)” in the sentence corresponding to the sensory evaluation. This modification is included in line 32 of the revised manuscript.

Comment 6 | References

References [95] and [96] appear to cite the same work (Taipe et al., 2022, Ciencia Tecnología Agropecuaria 23). This duplicate reduces the credibility of the literature review.
Requested action: Remove the duplicate reference and renumber all subsequent citations accordingly.

RESPONSE 6: We thank the reviewer for pointing out this possible duplication in the references.

After a detailed revision, it was confirmed that the indicated references do not correspond to the same work. In the revised version of the manuscript, these references were renumbered as [85] and [86], respectively, and correspond to different sources in terms of both type and content: one reference corresponds to a book on the characterization of soy-derived ingredients, whereas the other corresponds to a scientific article on Sacha Inchi by-products.

Nevertheless, the wording of the corresponding text (lines 654–663) was revised in order to improve precision and avoid possible ambiguities in the interpretation of the cited sources, retaining only the relevant references. Additionally, the consistency of the numbering of all citations throughout the revised manuscript was verified.

Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

This study developed a method for preparing a chicken nugget analogue using soy protein, and comprehensively analyzed its physicochemical properties, processing characteristics, and sensory evaluation. A comparative study was conducted between the prepared chicken nugget analogue, chicken nuggets, and plant-based nuggets, and the limitations of the research were also discussed. The paper is rich in research content and highly innovative in terms of the product. I suggest that it can be accepted for publication after addressing the following issues:

1. The number of references in the introduction is as high as 47, which is not typical for a regular research paper. It is recommended to reduce this appropriately to a reasonable number.

2. Table 1: The number of decimal places in the data should be kept consistent in writing.

3. In Table 1, the '2' in the molecular formula of water should be a subscript.

4. In the formulation optimization, the authors performed extensive work, including measuring various data. I suggest combining the images showing the appearance of the finished products made from different formulations (Figure 11) with these data, so that the differences between formulations can be presented more directly to readers.

5. Line 775: The temperature at which the water activity was measured should be provided.

6. The total number of references is as high as 131, which is somewhat excessive for a research paper.

Author Response

REVIEWER 2:

This study developed a method for preparing a chicken nugget analogue using soy protein, and comprehensively analyzed its physicochemical properties, processing characteristics, and sensory evaluation. A comparative study was conducted between the prepared chicken nugget analogue, chicken nuggets, and plant-based nuggets, and the limitations of the research were also discussed. The paper is rich in research content and highly innovative in terms of the product. I suggest that it can be accepted for publication after addressing the following issues:

First of all, thank you very much for all your appreciation of our manuscript by the reviewer. In “blue font”, you will find our answers to your major suggestions/queries about the research.

1. The number of references in the introduction is as high as 47, which is not typical for a regular research paper. It is recommended to reduce this appropriately to a reasonable number.

RESPONSE 1: We thank the reviewer for this observation.

In response to this observation, the introduction was carefully revised with the aim of reducing the number of references and improving the clarity and conciseness of the text while maintaining the necessary scientific support.

Redundant references within the same idea were removed, retaining only the most representative or recent ones. For example, in the general description of nuggets and meat analogues (lines 55–56), as well as in the applications of Sacha Inchi cake in bakery products (lines 68–69) and energy bars (line 71), the number of citations was reduced when multiple studies addressed the same content.

Likewise, in the description of applications in emulsified meat products (lines 71–73), only one representative reference supporting the use of Sacha Inchi oil and cake in these matrices was retained.

Additionally, the theoretical background related to the response variables (textural properties, cooking loss and cooking yield, and color parameters), together with their corresponding references (lines 92–101), was removed because these correspond to concepts that are already well established in food science literature.

In the experimental design section, a general reference related to mixture design types (lines 104–106) was removed because it was not essential to the focus of the study.

Finally, in the background paragraph related to studies on meat analogue nuggets (lines 111–114), the number of references was reduced, retaining only the most recent and relevant ones.

Overall, these modifications allowed a significant reduction in the number of references in the introduction, improving the conciseness of the manuscript without compromising its scientific rigor.

2. Table 1: The number of decimal places in the data should be kept consistent in writing.

RESPONSE 2: We thank the reviewer for this observation.

In response to this comment, Table 1 (line 146) was revised, and the number of decimal places was standardized across all reported data, ensuring consistency in the presentation (two decimal places) throughout the table. These corrections have been incorporated into the revised version of the manuscript.

3. In Table 1, the '2' in the molecular formula of water should be a subscript.

RESPONSE 3: We thank the reviewer for this observation.

In response to this observation, the chemical notation of water in Table 1 (lines 146–147) was corrected by using the appropriate subscript in the formula (H₂O). This correction has been incorporated into the revised version of the manuscript.

4. In the formulation optimization, the authors performed extensive work, including measuring various data. I suggest combining the images showing the appearance of the finished products made from different formulations (Figure 11) with these data, so that the differences between formulations can be presented more directly to readers.

RESPONSE 4: We thank the reviewer for this suggestion.

In response to this comment, Figure 11, which shows the appearance of the formulations (previously located in the methodology section, subsection “3.5.1. Mixture Design”, line 907), was relocated to the results section as Figure 1 (line 189), within the subsection “2.2. Analysis of the Nugget Formulations” (lines 182–192), immediately before the analysis of the physicochemical and textural variables.

This modification allows a better integration between the visual evaluation of the samples and the instrumental data, facilitating a more direct interpretation of the differences among formulations.

5. Line 775: The temperature at which the water activity was measured should be provided.

RESPONSE 5: We thank the reviewer for this observation.

In response to this observation, the measurement temperature for water activity (a_w) was incorporated into the corresponding section of the manuscript (current line 884), specifying that the measurements were performed at 25 °C.

6. The total number of references is as high as 131, which is somewhat excessive for a research paper.

RESPONSE 6: We thank the reviewer for this observation. In response to this comment, the manuscript bibliography was carefully reviewed. The number of references in the introductory section was reduced by removing redundant citations, decreasing the total number of references in the manuscript from 131 to 123.

Nevertheless, this number was considered appropriate because the study comprehensively addresses multiple stages of food product development, including the physicochemical characterization of raw materials, product formulation, mixture design, optimization, physicochemical characterization of the final product, and sensory analysis (including CATA evaluation).

Within this context, the broad scope of the study requires bibliographic support from different research areas, which justifies the number of references included. Furthermore, efforts were made to ensure that all references are relevant, up to date, and directly related to the content developed in the manuscript.