Chicken Heads as a Promising By-Product for Preparation of Food Gelatins

Round 1

Reviewer 1 Report

Title: Chicken heads as a promising by-product forpreparation of food gelatins

The reported manuscript descried a new and innovative biotechnological method for the production of gelatin from chicken heads a large by-products of food industries. In order to obtain gelatin, chicken by-product was processed by a proteolytic enzyme. Some factor affecting of gelatin amount and its properties were optimized by using an experimental design. In my opinion the manuscript is interesting and well written, I suggest the authors to make min or revision before publication in Molecules

Main comments:

How the authors have been selected the condition range (min and max) of each factor used into the experimental design?

Looking to figure 1 the maximum value of gelatin yield (%) was obtained at the maximum value of both amount of enzyme (%) and Extraction time (%). In the basis of this result are the author sure that increasing the range of these factors the gelatin yield did not increase too?

The same observation made above could be made for figure 2 are they sure that decreasing the range of the two factors (amount of enzyme (%) and Extraction time (%) the gelatin Bloom did not increase?

In my opinion reporting the analysis of variance graphically could be more immediate and intuitive to the reader than in a table

Author Response

Point-by-point responses to the reviewer comments.

Changes in the manuscript are highlighted in red colour.

Reviewer Comment: How the authors have been selected the condition range (min and max) of each factor used into the experimental design?

Response: The condition range (minimal and maximal values) of studied process factors were selected according to our previous knowledge. Our team have been focusing on processing of many poultry by-products. We published some papers on preparation of gelatins (and hydrolysates as well) from chicken feet and chicken skins (see below). Thus, the selection of appropriate condition range of process factors in this paper regarding processing chicken heads was easier.

Some of our previous papers:

Mokrejš, P.; Mrázek, P.; Gál, R.; Pavlačková, J. Biotechnological preparation of gelatines from chicken feet. Polymers 2019, 11 1060; doi: 10.3390/POLYM11061060.

Mokrejš, P.; Gál, R.; Janáčová, D. Plšková, M.; Brychtová, M. Chicken paws by-products as an alternative source of proteins. Oriental J. Chem. 2017, 33, 2209-2216; doi: https://doi.org/10.13005/ojc/330508.

Mokrejš, P.; Gál, R.; Pavlačková, J.; Janáčová, D.; Mrázek, P. Utilisation of chicken slaughterhouse collagen by-products for preparation of gelatines and hydrolysates. Chemical Papers 2019, 113, 121-125; doi: 10.1155/2015/247013.

Mrázek, P.; Mokrejš, P.; Gál, R.; Orsavová, J. Chicken skin gelatine as an alternative to pork and beef gelatines. Potravinarstvo Slovak J. Food Sci. 2019, 13, 224-233; doi: 10.5219/1022.

Reviewer Comment: Looking to figure 1 the maximum value of gelatin yield (%) was obtained at the maximum value of both amount of enzyme (%) and Extraction time (%). In the basis of this result are the author sure that increasing the range of these factors the gelatin yield did not increase too?

Response: Yes, we agree with your presumption. Increased amount of enzyme used in the pre-treatment of starting collagen material and increased extraction time will result in moderate increase of gelatin yield. On the other hand, gelatin quality (assessed mainly by the gelatin gel strength and viscosity) will decrease. At the maximal limits of studied process factors, i.e. 1.6 % of added enzyme and 4.0 h of extraction time the gelatin gel strength is approx. 110 Bloom and the viscosity is approx. 2.0 mPa.s. Depending on the application in food and pharmaceutical industry, gelatins with the gel strength within the range 100-300 Bloom and viscosity within the range 2.0-7.0 mPa.s are preferred. Thus, increasing the range of the factors is not favorable with respect to potential applications of prepared gelatins.

Reviewer Comment: The same observation made above could be made for figure 2 are they sure that decreasing the range of the two factors (amount of enzyme (%) and Extraction time (%) the gelatin Bloom did not increase?

Response: Yes, we agree with this conclusion as well. This fact was proved in our experiments as well. In the gelatin industrial practice the gelatin manufacturer has to balance between gelatin yield (the conversion of the starting collagenous material into gelatin) and gelatin characteristics – not only regarding gel strength and viscosity, but also other characteristics important for food- and pharmaceutical applications (see e.g. one of our previous papers). Thus, at industrial scale, the starting material is extracted more than one time: to prepare gelatins with very high Bloom value at reduced gelatin yield in the first extraction step, and, in the following extraction step(s) gelatins with lower Bloom value(s) with higher gelatin yield(s). In our study it was proved that second extraction (at higher extraction temperature) gives better total yield of extracted gelatins.

Reviewer Comment: In my opinion reporting the analysis of variance graphically could be more immediate and intuitive to the reader than in a table

Response: The table with the results of analysis of variance is now omitted. Instead of this, the results are presented in two ways. Firstly, the effects of studied process factors (and their interactions) on evaluated variables (gelatin yield, gelatin gel strength and gelatin viscosity) are presented by Pareto charts. Please, see changes in the manuscript in lines 117-120 (page 3) and on page 4 (lines up to 129) – Figure 1. Secondly, the results of p-values and regression equations (important values for the readers) are included in the text body – see chapter 2.1 (lines 131-132, 134-137), chapter 2.2 (lines 163-166) and chapter 2.3 (lines 187-189).

Note: Figures and Tables in the revised paper have been re-numbering.

Reviewer 2 Report

The authors studied the possibilities of the preparation of gelatin from chicken heads and technological factors influencing the yield and properties of the gelatins. The results showed that chicken heads can be a promising by-product for the preparation of gelatins for food, pharmaceutical and cosmetic industry. The reviewer wondering, how about the purity of the gelatins in this study, because the reviewer cannot see whether the authors quantified the amount of residual non-gelatin proteins in the products. 

Author Response

Point-by-point responses to the reviewer comments.

Changes in the manuscript are highlighted in red colour.

Reviewer Comment: The reviewer wondering, how about the purity of the gelatins in this study, because the reviewer cannot see whether the authors quantified the amount of residual non-gelatin proteins in the products. 

Response: The purity of prepared gelatins expressed by quantifying the residues of non-collagenous proteins was not assessed in this study. During the initial steps of the procedure, the starting collagen materials (after grinding and homogenization) is thoroughly purified in four steps with 0.1% NaOH and water. This treatment is sufficient for removal albumins, globulins and pigments. Many authors use the same procedure to remove unwanted non-collagen proteins from starting raw-materials. After that, the material is defatted with the act of solvents. Moreover, in the conditioning step (treating with protease) any eventually remained non-collagen globular proteins are hydrolyzed with the enzyme.

In our on-going research our team will focus on testing the suitability of chicken heads gelatins for some food applications (especially for production of gelatin gummies, marshmallows, and pies). Now, experimental works aimed on detail study of microbiological parameters of gelatins prepared under different technological conditions, including the influence of the methods of sterilization and drying (which may affect the microbiological purity of gelatins), is being carried out.