Characterization of Antioxidant Peptides from Thai Traditional Semi-Dried Fermented Catfish
, Tanutchaporn Rodsamai 1, Sawanya Charoenlappanit 2, Sittiruk Roytrakul 2, Atikorn Panya 3, Natthaporn Phonsatta 3, Ling-Zhi Cheong 4 and Worawan Panpipat 1,*Round 1
Reviewer 1 Report
This work present a characterization of the antioxidant peptides from a Thai traditional semi-dried fermented farmed catfish. I would like to congratulate the authors for the cientific rigor of the work.
I only have minor corrections to suggest.
Line 77-79 – letter type format is different.
Line 101-102: I think you want to say, that ‘’ all the centesimal composition analysis were performed according the AOAC guidelines.’’
Author Response
Reviewer 1
This work present a characterization of the antioxidant peptides from a Thai traditional semi-dried fermented farmed catfish. I would like to congratulate the authors for the cientific rigor of the work.
Ans: Thank you very much.
I only have minor corrections to suggest.
Line 77-79 – letter type format is different.
Ans: This is due to a technical error. We've already made the changes to the format.
Line 101-102: I think you want to say, that ‘’ all the centesimal composition analysis were performed according the AOAC guidelines.’’
Ans: It was changed to “All the centesimal composition analysis were performed according to the AOAC guidelines [19]. The Pla Duk Ra sample had 40.16±0.13% moisture, 35.95±0.60% protein, 10.86±0.45% fat, 7.71±0.24% ash, and 5.32±1.19% carbohydrate.”
Reviewer 2 Report
Type of manuscript: Article
Title: Characterization of Antioxidant Peptides from Thai Traditional Semi-Dried Fermented Catfish
Why did the author choose semi-dried fermented fish as the material?
What is the importance of semi-dried fermented fish?
What is the difference between dried and fermented fish and fermented fish?
Why does the test tube test for antioxidant determination represent the antioxidant capacity of the sample?
Why not carry out the antioxidant assay of the cell test?
Introduction
Line 47-54, Why does the author specifically explain the fermentation process of fish? Is this fermentation process semi-dry fermented fish or fermented fish?
Materials and Methods
Line 95-98, What is the storage method of Pla Duk Ra during transport to the laboratory? How is Pla Duk Ra made? Will there be differences in Pla Duk Ra purchased from different places?
Line 99, What are the setting conditions and temperature of the MK 5087M Panasonic Food Processor?
Line 109 and Line 111, What is the concentration of ethanol?
Line 109-110, To remove the proteins. How to remove protein?
Line 117, Why measure 280 and 420 nm?
Line 129, Why is the peptide content measured using the Bradford method instead of the Lowry method (Line 120)? What is the difference between the two methods?
Line 147-155, Why measure so many different antioxidant indicators? Why doesn't the author choose one of the antioxidant indicators with different characteristics for testing?
Line 156, Why only choose DPPH for measurement?
Line 176, What is the number of sample replicates for each experiment?
Results and Discussion
Line 184, Isn't it measuring 280 nm? Why is it the result of 294 nm?
Figure 1, How to distinguish between F1 and F2, and what is the basis? It should be explained in the material and method.
Line 199, F1 and F2 contained 19 and 14 amino acids. Why is the number of amino acids in F1 and F2 different? What are the possible reasons?
References
The journal name should be in italics.
Author Response
Reviewer 2
Title: Characterization of Antioxidant Peptides from Thai Traditional Semi-Dried Fermented Catfish
Why did the author choose semi-dried fermented fish as the material?
Ans: The reason for using semi-dried fermented catfish as a material was explained in the Introduction.
“Pla Duk Ra, a semi-dried fermented catfish, is one of Thailand's most popular traditional fish products, especially in the south…..”
“Fermented fish is a natural source of protein hydrolysate with a wide range of peptides with flavorful taste components and biological activity potential [4-5]…..”
What is the importance of semi-dried fermented fish?
Ans: The importance of semi-dried fermented catfish was originally described in the Introduction.
“Pla Duk Ra, a semi-dried fermented catfish, is one of Thailand's most popular traditional fish products, especially in the south. It is created by salting and fermenting the farmed hybrid catfish (Clarias macrocephalus ´ Clarias gariepinus), which is Thailand's most intensively farmed freshwater fish [1-2].”
“This product with high protein content has unique nutritional benefits for humans, and it also imparts delicacy with a strong excellent flavor developed during fermenta-tion [1].”
What is the difference between dried and fermented fish and fermented fish?
Ans: The difference in the final aw between dried fermented fish and semi-dried fermented fish is significant. Semi-dried foods, also known as intermediate moisture foods (IMF), are semi-moist foods (aw = 0.6-0.85), whereas dried foods have an aw of less than 0.6. In the Introduction, we said that the technique for producing semi-dried fermented catfish required sun drying until the aw of £ 0.85 was achieved.
“Finally, sun drying for 5 h was done till the aw of £ 0.85 was achieved, which was within the range of semi-dried food (aw = 0.6-0.85) [3].”
Why does the test tube test for antioxidant determination represent the antioxidant capacity of the sample?
Ans: In general, active substances can be screened for their antioxidant capacity using both in vitro and in vivo antioxidant tests. In this investigation, a variety of in vitro assays (free radical scavenging activities, reactive oxygen scavenging activities, and metal chelating activity) were used to determine the possible antioxidant mechanisms of the peptides.
Why not carry out the antioxidant assay of the cell test?
Ans: Thank you very much for your insightful suggestions. In the future, we will conduct an antioxidant assay for the cell test.
Introduction
Line 47-54, Why does the author specifically explain the fermentation process of fish? Is this fermentation process semi-dry fermented fish or fermented fish?
Ans: We presented the procedure for fermented fish in this paragraph because it can produce natural peptide during fermentation and semi-dried fermented fish can be produced later by drying the fermented fish. The semi-dried fermented catfish procedure was described in detail in the previous paragraph.
Materials and Methods
Line 95-98, What is the storage method of Pla Duk Ra during transport to the laboratory? How is Pla Duk Ra made? Will there be differences in Pla Duk Ra purchased from different places?
Ans: We originally stated that “Samples packed in plastic bags were carried to Walailak University's laboratory in a polystyrene foam box within 1 h.”.
The method for producing Pla Duk Ra was already explained in the Introduction. “According to the document of Thai Community Product Standard [3] and the observation from local producers in Southern Thailand, fresh fish was cleaned, headed, eviscerated, and drained for 3 h. The fish was then mixed with salt and sugar in a 10:0.4:0.4 (w/w/w) ratio and kept tightly sealed in earthen jar for 1 night at room temperature (27-29 °C) for fermentation. After that, the salted fish was sun-dried for 5 h. Then it was left to ferment for two days under the same conditions. The exudates are allowed to drip off during the fermenting process. Finally, sun drying for 5 h was done till the aw of £ 0.85 was achieved, which was within the range of semi-dried food (aw = 0.6-0.85) [3].”
We were worried about how Pla Duk Ra differed from place to place. So, throughout the experiment, we used the same lot of Pla Duk Ra from the same local producer.
Line 99, What are the setting conditions and temperature of the MK 5087M Panasonic Food Processor?
Ans: The details were added. “The flesh was manually separated from the bone upon arrival and mashed together using an MK 5087M Panasonic Food Processor (Selangor Darul Ehsan, Malaysia) at speed 1 for 10 min at room temperature to create a composite sample before peptide extraction.”
Line 109 and Line 111, What is the concentration of ethanol?
Ans: It was absolute ethanol.
Line 109-110, To remove the proteins. How to remove protein?
Ans: It was stated that “To remove the proteins, the mixture was maintained at 4°C for 20 h. Subsequently, the sample was centrifuged at 12,000 ´g (4 °C/ 20 min). The supernatant was filtered with a 0.45 mm Nylon filter after centrifugation under the same conditions, and ethanol was evaporated by an Eyela rotary evaporator (Model N-100, Tokyo, Japan).”
Line 117, Why measure 280 and 420 nm?
Ans: We already mentioned that “…….and the absorbance of the 1.0-mL fractions was measured at 280, and 420 nm using a UV-Vis spectrophotometer (UV-1900, Shimadzu, Kyoto, Japan) to monitor peptides, and Maillard reaction products (MRPs), respectively.”
Line 129, Why is the peptide content measured using the Bradford method instead of the Lowry method (Line 120)? What is the difference between the two methods?
Ans: When compared to the Lowry method, the Bradford method is more sensitive. The Lowry method was employed during fractionation. The peptide's concentration was diluted after successive purification using C18ZipTip. As a result, the Bradford approach, which is more sensitive, was used.
Line 147-155, Why measure so many different antioxidant indicators? Why doesn't the author choose one of the antioxidant indicators with different characteristics for testing?
Ans: In this investigation, a variety of in vitro assays (free radical scavenging activities, reactive oxygen scavenging activities, and metal chelating activity) were used to determine the possible antioxidant mechanisms of the peptides.
Line 156, Why only choose DPPH for measurement?
Ans: Because it was the most effective antioxidant mechanism, the DPPHâ—Ź scavenging activity was chosen to be studied in this section. This statement was added at the beginning of this paragraph.
Line 176, What is the number of sample replicates for each experiment?
Ans: The data were expressed as means ± standard deviations (SD) of three replications (n = 3) for all analyses.
Results and Discussion
Line 184, Isn't it measuring 280 nm? Why is it the result of 294 nm?
Ans: It was A280 and A420. The paragraph was revised “Figure 1a shows chromatograms acquired using Hitrap Sephadex-G25 columns. The peptide distribution was represented by A280, while the browning product was represented by A420. F1 with higher A420 was eluted first, followed by F2. Figure 1b depicts the appearance of CPE, F1, and F2. The A420 of 2.70±0.08, indicated the existence of MRPs in the CPE. Because of the reaction between amine-carbonyl compounds, MRPs can be produced during Pla Duk Ra fermentation. F1 (A420 = 0.07±0.00) was light yellow as well, although with a lesser intensity than CPE. The low intensity of A420 (0.05±0.00) made F2 the clearest. The higher molecular weight MRPs can be isolated sooner using gel filtration. As a result, they were predominantly found in the F1.”
Figure 1, How to distinguish between F1 and F2, and what is the basis? It should be explained in the material and method.
Ans: It was added in the Material and Method that “Due to differences in size distribution and browning intensity, two fractions were obtained: fraction 1 (F1) and fraction 2 (F2).” Also, it was noted in the Figure legend that “F1 represented the pooled fractions 3-10, while F2 represented the pooled fractions 11-18.”
Line 199, F1 and F2 contained 19 and 14 amino acids. Why is the number of amino acids in F1 and F2 different? What are the possible reasons?
Ans: The difference in the number of amino acids between F1 and F2 was attributable to the fractionation effect of gel filtration, which can partition peptides into fractions. This statement was added at the beginning of the paragraph.
References
The journal name should be in italics.
Ans: Done.
This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.
