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. 2022 Feb 3;17(2):e0263389. doi: 10.1371/journal.pone.0263389

Effect of collagen casing on the quality characteristics of fermented sausage

Xinlei Yan 1,#, Le Yang 1,#, Yanni Zhang 1, Wenying Han 1, Yan Duan 1,*
Editor: A M Abd El-Aty2
PMCID: PMC8812867  PMID: 35113961

Abstract

Objective

Fermented sausage is popular all over the world for its rich nutrition and unique flavor. Sausage casing is one of the key factors affecting the quality of fermented sausage. However, there is little information involved in this field.

Methods

In this study, collagen casings were used as a wrapping material, and natural casings (pig casings) were used as a control. The effects of the two types of casings on biogenic amine content and other quality characteristics of fermented sausage were analyzed with increasing the storage time.

Results

The results showed that with storage time increasing, the hardness and gumminess of fermented sausage in collagen casing (CC) group were higher than those in pig casing (PC) group (P<0.05), while the elasticity in CC group was lower than that in PC group (P<0.05). In the processing and storage period, there was no significant difference in the type and content of flavor substances between the two groups. More importantly, the contents of tryptamine, putrescine, cadaverine, histamine, tyramine and phenyethylamine in fermented sausage of CC group were lower than those in PC group (P<0.05).

Conclusion

In conclusion, this study revealed that CC could improve the quality characteristics of fermented sausage and reduce the content of biogenic amines in fermented sausage, which provides a theoretical basis for the choice of casings in industrial production in the future.

Introduction

Fermented sausage is one of the common fermented meat products. Due to its unique flavor and taste, it is favored by worldwide consumers [1]. It is usually stored and transported at normal temperature and can be eaten directly without cooking. However, many safety risks occur during the processing and storage of fermented sausages, such as oxidative rancidity, pathogenic bacteria and an excessive content of biological amines [2].

Proteins in meat could be converted into biogenic amines under the action of amino acid decarboxylase secreted by microorganisms in the fermentation process [3]. Meanwhile, biogenic amines are not volatile and have high thermal stability. Thus, the biogenic amine content is a common phenomenon in fermented sausages. A certain amount of biogenic amines is indispensable for human beings, but they are potentially toxic to the human body and even cause obvious adverse reactions when an individual ingests excessively. Therefore, the biogenic amine content is regarded as an important indicator for the safety assessment of fermented meat products, and it has been attracting attention from an increasing number of related research groups [4].

Casings play an important role in the industrial production of fermented sausage [5]. There are a variety of casings with different functional characteristics in market at present according to the different demands for meat packaging material in different countries and regions. Casings can be further divided into natural casings and collagen casings in terms of edibility. Natural casings are made from the fresh intestines of healthy animals after some processing, such as pig and sheep [9]. But it cannot be adapted to large-scale industrialized mass production due to their single source and poor processing performance. Collagen casing is an edible artificial casing, which is made from animal dermis. It has stable performance during industrial filling, which can produce products of uniform specifications, reduce the generation of defective products and achieve maximum benefits [6–10].

The maturation process of fermented sausage is easily affected by the type of sausage casing. At present, most studies focus on the comparison of structural properties between collagen casings and natural casings [11, 12]. However, there is little information about the effects of collagen casing on fermented sausage quality characteristics [2, 5, 13]. Serio et al. showed that different types of casings had a great influence on fermented sausages quality, which may affect biogenic amine content and the production of some volatile flavor compounds through affecting proteolysis in ripening time [6]. Škaljac et al. indicated that at the end of drying as well as at the end of the storage period, the total content of 13 US-EPA PAHs (U.S. Environmental Protection Agency Polycyclic Aromatic Hydrocarbons) was lower in sausages with collagen casings than in sausages with natural casings [14]. Zając et al. showed that sausages in the collagen casing were softer and had higher a* and b* color parameters than those with ovine casings [15].

In this study, two types casings, collagen casings (CC) and natural casings (pig casings, PC), were investigated, and the biogenic amine contents and other quality characteristics of fermented sausage were determined. Our results may provide theoretical evidence for the production of fermented sausage and the selection of its casing types (collagen casing or natural casing) from a security perspective.

Materials and methods

Preparation of the fermented sausages

All meat was taken from Sunite sheep that was purchased from Xilin Gol League, Inner Mongolia, China and ingredients were purchased from a supermarket (East tile kiln, Saihan District, Hohhot, Inner Mongolia, China). The ingredients in traditional formulation (salt 2.5%, Inner Mongolia Ejinur Salt Co., Ltd; sodium nitrate 100 mg/kg; sodium nitrite 70 mg/kg, Sichuan Jinshan Pharmaceutical Co., Ltd, Sichuan, China; glucose 0.5%, Jining Huipeng Chemical Co., Ltd, Jining, Shandong, China; black pepper 0.5% and sucrose 0.5%, Shandong Tuohao Food Co., Ltd, Dezhou, Shandong, China) were added to a mixture of 80% chopped sheep hind leg and 20% sheep tail fat. All fermented sausages were inoculated with a recommended commercial starter culture composed of Pediococcus pentosaceus and Staphylococcus xylosus (F-1 BactofermTM, Chr-Hansen, Denmark) for traditionally fermented meat products, with an amount of 25 g/200 kg added. The production of fermented sausage was divided into two groups according to the experimental design. The fermented sausages in one group were made by pig casing as packing material (PC group, casing diameter was 22 mm which were purchased from Shunping Qianyuan Electronic Commerce Co., Ltd). And the other group was made by collagen casing as the packing material (CC group, casing diameter was 22 mm, Shunping Qianyuan Electronic Commerce Co., Ltd, Baoding, Hebei, China).

Mutton was chopped up by chopping machine (ZB-25, Zhucheng Holy land food packaging machinery Factory, Weifang, Shandong, China, chopping speed, 1800 rpm; inject rate, 200 rpm) and salted with auxiliary materials (1 day). After curing, fermented sausages were formed by sausage enema mechanism (JQC-Ⅱ, Zhucheng Holy land food packaging machinery factory, Weifang, Shandong, China, filling speed, 100–500 kg/h and error was less than 5 grams).The stuffed sausages with a diameter of 10 cm were hung in a box with constant temperature and humidity (RH). Fermentation was performed at 25°C and 95% relative humidity for the first 3 days. For the ripening process, the temperature was set at 16°C and 90%~70% RH for 14 days. Samples were taken on day 0 (after salting), day 3 (after fermentation), day 11 (after processing), day 21 (after storage for 10 d), day 41 (after storage for 30 d) and day 61 (after storage for 50 d). The parallel test was carried out 3 times (3 batches of sausages were made at the same time), and 3 sausages were randomly selected from each batch at a certain time to determine the relevant indicators.

Physical and chemical analysis

Color measurement

Color measurement of sausage was carried out with TCP2 automatic color meter (Beijing Xin’ao Yike Photoelectricity Technology Co., Ltd, Beijing, China), which referred to the method of Zheng et al. [16]. The sausage products were cut into the slices about 0.5 cm thick and each sample was repeated 3 times individually. The color meter was preheated for 15 min and calibrated with standard plates Y = 87.3, X = 0.3158 and Y = 0.3233 before detection. The instrument according to the sample spot area: φ 20 mm. The standard light source for color measurement was D65 light source. The results were expressed in the CLE (L*, luminance value; a*, redness value; and b*, yellowness value) system.

Water activity analysis

The sample was shredded and placed in a vessel (Wuxi Huake Instrument & Meter Co. Ltd, Wuxi, Jiangsu, China), and the water activity was measured using an HD-3A water activity meter (Wuxi Huake Instrument & Meter Co. Ltd, Wuxi, Jiangsu, China). Sample was chopped and putted it into a sample plate (capacity: 20 mL).

pH value analysis

The shredded sample (2 g) was homogenized in 18 ml of normal saline (0.85% NaCl, v/v), and the pH value of the homogenized sample was determined using a PB-10 pH meter (Sartorius, AG, Germany). The pH meter was calibrated with standard solution (pH 4.0 and 7.0).

Microbiological analysis

The samples were gradually diluted, poured into a plate and were incubated at 37°C for 24 h in a LRH-150F biochemical incubator (Shanghai Yiheng Technology Co., Ltd, Shanghai, China). The number of lactic acid bacteria was cultured and counted with MRS (man rogosa sharpe) medium (Nanjing Jianceng Biotechnology Co., Ltd, Nanjing, China, dilution solution: distilled water), and the total aerobic mesophilic bacteria cultured and counted with PCA (plate counting agar) medium (Nanjing Jianceng Biotechnology Co., Ltd, Nanjing, China, dilution solution: distilled water). The appropriate gradient plate (colony count ranged from 30 to 300) was selected for counting, and the average number of colonies on the same dilution plate was calculated multiplied by the corresponding dilution multiple as the colony result per milliliter of sample.

Texture profile analysis (TPA)

The texture profile was analyzed by a texture analyzer (QTS texture instrument: FTC Corporation, USA). The determination parameters were set as follows: pre-test rate, 2mm/s; pilot test rate, 2mm/s; post-test rate, 2mm/s; test interval, 5s; compression ratio, 50%; trigger force, 5g; trigger type, automatic. The center of the samples was removed and then cut into cubes of 1 cm×1 cm×1 cm. The hardness, elasticity, adhesiveness, gumminess and chewiness were measured by two cycles at room temperature by using a "P-100" probe [17].

Volatile compound analysis

The volatile compounds of fermented sausages were extracted using HS-SPME and analysed using a GC-MS system (ISQ GC-MS instrument, Thermo Fisher Scientific, USA). The extraction was referred to the method proposed by Luo et al. [18] with some modifications. The extraction head (SPME extractor, Supelco, USA) was inserted into the sample flask with a distance of 1 cm from the sample, adsorbed at 60°C for 40 min. And thenthe extraction head was inserted into the injection port (250°C) of the GC instrument (ISQ GC-MS instrument, Thermo Fisher Scientific, USA) to desorb the analytes for 3 min. All samples were extracted in triplicate.

Determination of biogenic amines

The biogenic amine content was tested by using an Intelligent HPLC (High-performance liquid chromatograph, Agilent Technologies Inc., California, USA) following the previous method report [19]. The determination was performed on C18 column (250mm in length, 4.6mm in inner diameter, packing particle size of 5μm) under the condition of that UV detection wavelength, 254 nm and column temperature, 35°C and the sample size was 20 μL. Mobile phase A was 90% acetonitrile and 10% 0.01mol/L ammonium acetate solution containing 0.1% acetic acid, mobile phase B was 10% acetonitrile and 90% 0.01mol/L ammonium acetate solution containing 0.1% acetic acid. The flow rate of the sample was 0.8 mL/min. The details of gradient elution procedure was shown in Table 1.

Table 1. Liquid phase gradient elution procedure.

Time (min) 0 22 25 32 32.01 37
Mobile phase A/% 60 85 100 100 60 60
Mobile phase B/% 40 15 0 0 40 40
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The bioamine standard curve was plotted by taking tryptamine, phenylethylamine, putrescine, cadaverine, tyramine, spermidine and histamine standard solution concentrations as the horizontal coordinate and sampling the standard solution concentrations from 0.1, 0.25, 0.5, 1, 1.5, 2.5 and 5 μg/mL in sequence, and the ratio of the obtained peak area to the internal standard peak area as the vertical coordinate.

Statistical analysis

The quality data of fermented sausage were summarized using descriptive statistics. The L*, a*, b*, pH and aw of fermented sausage in the two groups were measured at the processing and storage time points of 0 d, 3 d, 11 d, 21d, 41d and 61d. Biogenic amine content, TPA and volatile flavour composition of fermented sausage in the two groups were measured at the processing and storage time points of 11 d, 21 d, 41 d and 61d. Random terms were included fermentation humidity and ambient temperature. Approximate one-way ANOVA and Tukey’s post hoc test for each fixed effect were conducted and critical value for a statistically important effect taken at P < 0.05. Statistical analyses were performed using statistics 23.0 software (SPSS, USA). Three technical replicates were measured at each time per treatment, and the data are expressed as the average ± standard deviation (S.D.).

Results

Physical and chemical analysis

The results showed that the pH of both the groups decreased first and then increased and reached the lowest value on the third day, indicating the end of fermentation. As shown in Table 2, the pH value of CC group was significantly lower than that of PC group at each period after 3 d of fermentation (P<0.05). The aw values of the two groups remained stable after decreasing but showed no significant difference (P>0.05). The values were kept below 0.8 since the end of the dry period (11 d). However, the results showed that L* in both groups presented the same tendency to level off after decline, and it was significantly higher at 3 d than that at storage period (P <0.05). a* and b* showed a fluctuating upward trend, and the value at 61 d was highest (P <0.05).

Table 2. The effect of the casing type on physical and chemical properties of fermented sausage.

Group Processing and storage time
0 d 3 d 11 d 21 d 41 d 61 d
L* PC 44.3±0.3Da 44.0±0.3Ca 36.0±0.6Aa 36.1±0.4Aa 36.0±0.08Aa 39.3±0.6Ba
CC 44.2±0.07Ca 43.1±0.4Ba 34.0±0.1Aa 35.0±0.5Aa 34.6±0.3Ab 34.0±0.3Ab
a* PC 20.6±0.2Bab 20.1±0.2Aa 24.3±0.07Ca 21.4±1.2Ca 21.3±0.2BCa 24.5±0.04CDa
CC 21.2±0.1ABa 21.1±0.08Ba 22.0±0.9Ba 19.8±0.2Aa 20.0±0.1Ab 22.1±0.2Bb
b* PC 9.0±0.04Ba 9.0±0.1ABa 10.1±0.1Ba 8.0±0.7Aa 8.2±0.3Ba 9.7±0.1Ba
CC 9.1±0.01Ba 8.9±0.04Ba 8.8±0.3Bb 7.7±0.09Aa 7.8±0.2Aa 9.0±0.3Ba
pH PC 5.9±0.02Aa 5.1±0.04Ba 5.8±0.02Aa 5.7±0.003Aa 5.7±0.01Aa 5.9±0.01Aa
CC 6.0±0.13Aa 5.0±0.02Ba 5.1±0.01Bb 5.4±0.02Cb 5.5±0.006Db 5.3±0.02Cb
aw PC 0.9±0.001Aa 0.9±0.003Ba 0.8±0.003Ca 0.7±0.002Da 0.7±0.001Ea 0.7±0.003Ea
CC 0.9±0.001Aa 0.9±0.001Ba 0.8±0.003Cb 0.7±0.001Db 0.8±0.001Ea 0.7±0.003Fb
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Data in the table were the mean ± S.D. of the three fermented sausage per experimental group of three independent experiments. Different capital letters indicated significant differences over time within the same group (P<0.05) and different lowercase letters indicated significant differences between groups at the same time (P<0.05), while the same letter indicated no significant difference (P>0.05). PC: pig casing, CC: collagen casing; L*: luminance value; a*: redness value; b*: yellowness value; aw: water activity.

Microorganism analysis

The number of lactic acid bacteria (Table 3) and the total aerobic mesophilic bacteria (Table 4) of fermented sausages in both groups showed a consistent trend over time, which appeared from 0 to 3 d after a rapid increase in fluctuations. However, there was no significant difference between CC and PC groups (P>0.05).

Table 3. The change in lactic acid bacteria number during processing and storage period.

Time (d) 0d 3d 11d 21d 41d 61d
PC 5.962±0.014Aa 8.081±0.076Da 8.151±0.041Da 7.291±0.033Ba 7.987±0.092Da 7.751±0.016Ca
CC 6.326±0.032Ab 7.974±0.048Da 8.335±0.039Eb 7.463±0.095Bb 7.697±0.068Cb 7.504±0.013Bb
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Data in the table were the mean ± S.D. of three fermented sausage per experimental group of three independent experiments. Different capital letters indicated significant differences over time within the same group (P<0.05) and different lowercase letters indicated significant differences between groups at the same time (P<0.05), while the same letter indicated no significant difference (P>0.05).

Table 4. Changes of viable bacteria number.

Time (d) 0d 3d 11d 21d 41d 61d
PC 6.756±0.196Aa 8.189±0.114Da 7.881±0.055Ca 7.049±0.157ABa 7.264±0.186Ba 7.777±0.109Ca
CC 6.366±0.021Aa 7.817±0.178CDb 8.079±0.234Da 7.408±0.111Bb 7.233±0.167Ba 7.497±0.287BCa
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Data in the table were the mean ± S.D. of three fermented sausage per experimental group of three independent experiments. Different capital letters indicated significant differences over time within the same group (P<0.05) and different lowercase letters indicated significant differences between groups at the same time (P<0.05), while the same letter indicated no significant difference (P>0.05).

Texture index analysis

Texture quality is an important indicator of the chew ability and taste of meat products, which could determine the quality of sausage. The results showed that hardness of fermented sausage tended to increase with increasing storage time, and hardness value of CC group was significantly higher than that of PC group (P<0.05). The gumminess in CC group was significantly higher than that in PC group at 11, 41 and 61 d (P<0.05), meanwhile the chewiness in CC group was significantly higher than that in PC group (P<0.05) (Table 5).

Table 5. Changes of TPA during processing and storage period.

Characteristic index Group 11 d 21 d 41 d 61 d
Hardness/g PC 1042.0±270.4Aa 3378.9±117.4Ba 2884.2±454.7Ba 3258.5±573.8Ba
CC 2668.9±166.9Ab 2783.1±248.9Aa 4228.5±124.5Bb 4531.5±72.1Ba
Elasticity/mm PC 0.6±0.04Aa 0.5±0.002Ba 0.6±0.05Aa 0.6±0.005ABa
CC 0.5±0.008ABa 0.6±0.008ABb 0.6±0.01Bb 0.5±0.01Ab
Adhesiveness/g.sec PC 0.5±0.02Aa 0.5±0.008Aa 0.5±0.02Aa 0.5±0.02Aa
CC 0.5±0.01Aa 0.5±0.007Ba 0.5±0.006ABa 0.5±0.01Ba
Gumminess/g PC 911.8±123.2Aa 1561.6±40.08Aa 1430.6±205.8Aa 1585.0±337.8Aa
CC 1407.6±56.6Ab 1407.9±167.7Aa 1999.7±154.6Ba 2189.6±43.4Ba
Chewiness/g·mm PC 585.0±100.9Aa 698.9±62.7ABa 935.0±218.5ABa 1073.0±88.8Ba
CC 782.4±16.3Aa 730.8±63.1Aa 1153.5±79.1Ba 1156.3±57.9Ba
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Data in the table were the mean ± S.D. of the three fermented sausage per experimental group of three independent experiments. Different capital letters indicated significant differences over time within the same group (P<0.05) and different lowercase letters indicated significant differences between groups at the same time (P<0.05), while the same letter indicated no significant difference (P>0.05).

Volatile flavour composition of sausages

114 volatile flavor compounds were detected in fermented sausages during processing and storage time, including 20 alcohols, 16 aldehydes, 15 acids, 11 esters, 2 phenols, 3 ketone, 39 terpenes and 8 other compounds (Table 6). With the increase of processing and storage time, the flavor substances in fermented sausage gradually decreased. The contents of terpenes in the two groups accounted for about 1/3 of the total contents. The contents of Acetoin (3-hydroxy-2-butanone) in PC group was significantly higher than that in CC group (P<0.05) at 11d and 41d.

Table 6. Volatile flavour compounds identified and quantified (μg/100g) in fermented sausages by GC–MS during storage.

Flavor Compounds RT Formula Contents (μg/100g)
11d 21d 41d 61d
CC PC CC PC CC PC CC PC
Alcohols (20)
Ethanol 1.34 C2H6O 0.48±0.12a 0.45±0.16a 1.09±0.17 Nd Nd 1.16±0.07 Nd Nd
1-Butanol, 3-methyl- 4.02 C5H12O 0.11±0.03a 0.12±0.03a Nd Nd Nd Nd Nd Nd
2,3-Butanediol 5.33 C4H10O2 1.07±0.33a 1.48±0.33a 1.32±0.14a 1.63±0.53a 1.38±1.01a 2.38±0.19a 1.73±0.38a 1.57±0.07a
1-Hexanol 7.71 C6H14O 0.06±0.01a 0.08±0.02a Nd Nd Nd Nd Nd Nd
2-Heptanol 8.61 C7H16O 0.04±0.02a 0.04±0.02a Nd Nd Nd Nd Nd Nd
1,6-Octadien-3-ol, 3,7-dimethyl- 14.15 C10H18O 0.54±0.01a 0.54±0.14a 0.92±0.02a 1±0.26a 0.95±0.33b 1.35±0.07a 0.87±0.14a 0.66±0.1a
endo-Borneol 16.15 C10H18O 0.23±0.01a 0.2±0.05a 0.33±0.01a 0.39±0.14a 0.46±0.21a 0.51±0.03a 0.38±0.07a 0.3±0.05a
Terpinen-4-ol 16.33 C10H18O 0.11±0.01a 0.12±0.03a 0.17±0.01a 0.22±0.09a Nd 0.27±0.02 0.21±0.03a 0.16±0.02b
α-Terpineol 16.72 C10H18O 0.18±0.02a 0.2±0.05a 0.36±0.02a 0.37±0.11a 0.41±0.15a 0.53±0.05a 0.32±0.04a 0.28±0.05a
Geraniol 17.97 C10H18O 0.1±0.01a 0.11±0.03a Nd Nd Nd Nd Nd Nd
1-Pentanol 4.83 C5H12O 0.12±0.02b 0.12±0.04a 0.54±0.01a 0.62±0.15a 0.46±0.16b 0.7±0.06a 0.41±0.06a 0.31±0.05b
R-(-)-1,2-propanediol 7.23 C3H8O2 Nd 0.23±0.21 Nd Nd Nd Nd Nd Nd
2,5-Hexanediol 9.85 C6H14O2 Nd 0.03±0.01 Nd Nd Nd Nd Nd Nd
Ethanolamine 2.74 C2H7NO Nd Nd 0.67±0.05a 0.13±0.03b 0.27±0.07 Nd 0.43±0.07a 0.39±0.06a
Diethylene glycol 3.42 C4H10O3 Nd Nd 6.07±0.11a 6.41±1.84a 2.79±0.83b 5.89±0.47a 3.03±0.63a 2.6±0.36a
1-Octen-3-ol 10.89 C8H16O Nd Nd 0.42±0.01 Nd Nd Nd 0.61±0.11 Nd
Isoborneol 16.16 C10H18O Nd Nd 0.33±0.01a 0.39±0.14a Nd Nd Nd Nd
3-Methyl-1-butanol 1.44 C5H12O Nd Nd Nd 1.03±0.14 Nd 1.33±0.09 0.62±0.09b 2.1±0.19a
2-Butanol, 3-methyl- 5.34 C5H12O Nd Nd Nd Nd 1.3±0.49 Nd Nd Nd
3-Cyclohexen-1-ol, 4-methyl-1-(1-methylethyl)-, (R)- 16.33 C10H18O Nd Nd Nd Nd 0.24±0.13 Nd Nd Nd
Aldehydes (16)
Pentanal 3.17 C5H10O 0.56±0.13a 0.43±0.12a 0.81±0.08a 0.84±0.13a 0.35±0.12b 0.54±0.03a 0.37±0.05a 0.34±0.01a
Hexanal 5.68 C6H12O 0.42±0.07a 0.43±0.11a 5.22±0.4a 6.27±1.2a 1.16±0.47b 4.07±0.31a 1.87±0.33a 1.16±0.14b
Heptanal 8.65 C7H14O 0.48±0.04a 0.43±0.1a 2.38±0.06a 2.79±0.67a 1.03±0.3b 1.69±0.08a Nd Nd
Benzaldehyde 10.56 C7H6O 0.49±0.11a 0.55±0.12a 1.69±0.06a 1.51±0.49a 1.56±0.57b 2.73±0.24a 1.08±0.14a 0.91±0.15a
Benzeneacetaldehyde 12.88 C8H8O 0.12±0.01a 0.15±0.03a 0.72±0.01a 0.93±0.24a 0.59±0.24a 0.88±0.11a 0.62±0.07a 0.52±0.12a
Nonanal 14.33 C9H18O 1.13±0.29a 1.21±0.34a 2.5±0.04b 3.57±0.71a 1.6±0.53b 3.38±0.26a 3.45±0.81a 2.93±0.26a
2-Heptenal, (E)- 10.33 C7H12O Nd Nd 0.33±0.02a 0.42±0.15a Nd Nd Nd Nd
Octanal 11.58 C8H16O Nd Nd 2.92±0.09a 3.79±0.94a Nd Nd Nd Nd
2-Octenal, (E)- 13.16 C8H14O Nd Nd 0.5±0.09a 0.51±0.13a Nd Nd Nd Nd
2-Nonenal 15.82 C9H16O Nd Nd 0.63±0.05a 0.69±0.19a Nd Nd Nd Nd
2-Decenal, (E)- 18.32 C10H18O Nd Nd 0.22±0.03a 0.32±0.09a Nd 0.39±0.03 Nd Nd
Butanal, 3-methyl- 3.17 C5H10O Nd Nd Nd 0.84±0.13 Nd 0.54±0.03 0.44±0.15a 0.3±0.02a
Decanal 16.92 C10H20O Nd Nd Nd 0.28±0.1 Nd Nd Nd 0.14±0.04
Pentadecanal- 27.11 C15H30O Nd Nd Nd 0.38±0.01 0.42±0.19 Nd 0.36±0.06a 0.34±0.07a
Tetradecanal 27.11 C14H28O Nd Nd Nd 0.38±0.01 0.42±0.19a 0.54±0.06a 0.36±0.06a 0.34±0.07a
2-Nonenal, (E)- 15.82 C9H16O Nd Nd Nd Nd 0.19±0.06b 0.43±0.07a Nd 0.21±0.06
Acids (15)
Ala-Gly 1.18 C5H10N2O3 0.37±0.05a 0.21±0.16a Nd Nd Nd Nd Nd Nd
Malonamic acid 1.28 C3H5NO3 0.31±0.13a 0.33±0.15a 0.83±0.07a 0.63±0.16a 0.28±0.11b 0.75±0.07a 0.44±0.07a 0.4±0.04a
Glycine 1.62 C2H5NO2 1.61±0.62a 2.48±0.86a Nd Nd Nd 1.97±0.2 Nd Nd
Acetic acid 1.93 C2H4O2 5.81±1.79a 8.53±1.57a 9.63±0.24a 10.26±3.68a 9.37±3.4b 15.49±1.08a Nd 9.35±0.89
Methylmalonic acid 1.28 C4H6O4 5.83±1.40 Nd 6.07±0.11a 6.41±1.84a 2.79±0.83b 5.89±0.47a 3.03±0.63a 2.6±0.36a
Propanoic acid, 2-methyl- 4.47 C4H8O2 0.05±0.02a 0.08±0.03a Nd Nd Nd Nd Nd Nd
Butanoic acid, 3-methyl- 6.96 C5H10O2 0.27±0.09b 0.5±0.13a Nd Nd Nd Nd Nd 0.34±0.06
Butanoic acid, 2-methyl- 7.17 C5H10O2 0.11±0.02a 0.16±0.04a Nd Nd Nd Nd Nd Nd
Hexanoic acid 10.89 C6H12O2 0.27±0.01a 0.28±0.23a 0.95±0.04a 0.9±0.34a 0.54±0.3b 1.27±0.2a 0.83±0.14a 0.66±0.13a
Octanoic acid 16 C8H16O2 0.43±0.07a 0.59±0.16a 1.12±0.05a 1.71±0.64a 1.35±0.66b 3.43±0.28a 1.61±0.38b 2.65±0.45a
n-Decanoic acid 20.6 C10H20O2 Nd Nd 0.33±0.01a 0.36±0.16a 0.62±0.29a 0.53±0.04a 0.66±0.09a 0.62±0.14a
n-Hexadecanoic acid 28.36 C16H32O2 Nd Nd 0.69±0.11 Nd Nd Nd Nd Nd
Octadecanoic acid 28.36 C18H36O2 Nd Nd 0.69±0.11 Nd Nd Nd Nd Nd
Pentanoic acid 10.95 C5H10O2 Nd Nd Nd 0.9±0.34 0.54±0.3 Nd 0.83±0.14 Nd
N-Acetyl-L-alanine 1.26 C5H9NO3 Nd Nd Nd Nd 0.28±0.11b 0.75±0.07a Nd Nd
Esters (11)
Acetic acid, anhydride with formic acid 2.04 C3H4O3 0.02±0.00a 0.02±0.01a 0.1±0.02a 0.03±0.02b 0.04±0.01a 0.05±0.01a Nd Nd
Propanoic acid, 2-hydroxy-, ethyl ester, (S)- 5.99 C5H10O3 0.2±0.02a 0.16±0.04a Nd Nd Nd Nd Nd Nd
2-Heptanol, acetate 12.44 C9H18O2 0.08±0.01a 0.08±0.02a Nd Nd Nd Nd Nd Nd
Octanoic acid, ethyl ester 16.57 C10H20O2 0.15±0.03a 0.15±0.05a Nd 0.53±0.15 0.44±0.16b 0.9±0.06a 0.44±0.08a 0.46±0.01a
Bornyl acetate 18.77 C12H20O2 0.06±0.01a 0.07±0.02a Nd Nd Nd Nd Nd Nd
Acetic acid, oxo-, methyl ester 2.03 C3H4O3 Nd Nd 0.03±0.005a Nd Nd Nd Nd Nd
n-Caproic acid vinyl ester 11.07 C8H14O2 Nd Nd 1.75±0.07 1.72±0.46 Nd Nd Nd Nd
Octanoic acid, methyl ester 14.76 C9H18O2 Nd Nd Nd Nd Nd 0.34±0.04 Nd 0.2±0.02
Butyrolactone 9.22 C4H6O2 Nd Nd Nd Nd Nd Nd 0.42±0.07a 0.32±0.02b
Butanoic acid, 4-hydroxy- 9.22 C4H8O3 Nd Nd Nd Nd Nd Nd 0.42±0.07a 0.32±0.02b
Decanoic acid, ethyl ester 21.21 C12H24O2 Nd Nd Nd Nd Nd Nd Nd 0.73±0.95
Phenols (2)
Phenol 3.09 C6H6O 0.31±0.09a 0.31±0.08a Nd Nd Nd Nd 0.6±0.08a 0.47±0.05b
Eugenol 20.48 C10H12O2 0.24±0.04a 0.28±0.06a 0.38±0.05a 0.39±0.13a 0.66±0.27a 0.83±0.07a 0.5±0.07a 0.48±0.07a
Ketones (3)
Acetoin 3.43 C4H8O2 7.11±3.62b 13.26±3.46a 6.07±0.11a 6.41±1.84a 2.79±0.83b 5.89±0.47a 3.03±0.63a 2.6±0.36a
2,3-Octanedione 11.07 C8H14O2 Nd Nd 1.75±0.07 Nd Nd Nd Nd Nd
5-Hepten-2-one, 6-methyl- 11.05 C8H14O Nd Nd Nd Nd Nd Nd Nd 0.4±0.07
Terpenes (39)
Cyclobutene, 2-propenylidene- 4.7 C7H8 0.09±0.01a 0.08±0.02a Nd Nd Nd Nd Nd Nd
α-Pinene 9.34 C10H16 0.2±0.03a 0.2±0.06a Nd Nd Nd Nd Nd Nd
Camphene 9.86 C10H16 0.22±0.04a 0.27±0.12a 0.27±0.05 Nd 0.31±0.15a 0.35±0.04a 0.25±0.04a 0.17±0.05b
Bicyclo[3.1.1]heptane, 6,6-dimethyl-2-methylene-, (1S)- 10.69 C10H16 0.21±0.05a 0.67±0.63a Nd Nd 1.35±1.07a 0.28±0.03ab Nd 0.76±0.54
β-Pinene 10.69 C10H16 0.21±0.05a 0.19±0.06a Nd Nd Nd Nd Nd Nd
β-Ocimene 11.98 C10H16 2.07±0.28 Nd Nd Nd Nd Nd Nd Nd
2-Carene 13.72 C10H16 0.07±0.07a 0.04±0.01a Nd Nd Nd Nd Nd Nd
Limonene 12.18 C10H16 2.96±0.38a 3.03±0.87a 2.68±0.03a 2.78±0.84a 3.71±1.51a 2.49±2.13ab 2.55±0.4a 1.96±0.16b
β-Phellandrene 12.24 C10H16 3.49±0.34a 3.8±0.96a 2.7±0.07a 2.74±0.81a 3.86±1.56a 4.43±0.53a 2.85±0.42a 2.2±0.19b
Bicyclo[3.1.0]hexane, 4-methylene-1-(1-methylethyl)- 10.69 C10H16 0.21±0.05 Nd 2.7±0.07a 2.74±0.81a 3.86±1.56a 4.43±0.53a 2.85±0.42a 2.2±0.19b
Cyclohexene, 4-methylene-1-(1-methylethyl)- 12.24 C10H16 3.49±0.34 Nd 2.7±0.07a 2.74±0.81a 3.86±1.56a 4.43±0.53a Nd 2.2±0.19
Eucalyptol 12.28 C10H18O 0.25±0.02b 0.11±0.06a 0.18±0.01a 0.14±0.03b 0.18±0.06a 0.19±0.01a 0.14±0.02a 0.03±0.02b
γ-Terpinene 13 C10H16 0.15±0.02a 0.15±0.04a Nd Nd 0.2±0.08 Nd Nd Nd
Cyclohexene, 1-methyl-4-(1-methylethylidene)- 11.83 C10H16 0.07±0.04a 0.13±0.09ab Nd Nd Nd Nd Nd 1.96±0.16
Camphor 15.53 C10H16O 0.09±0.01a 0.1±0.02a Nd Nd Nd Nd Nd Nd
Anethole 18.99 C10H12O 2.47±0.21a 2.79±0.69a 4.46±0.08a 4.85±1.42a 5.89±2.35a 7±0.55a 4.41±0.64a 4.03±0.62a
Cyclohexene, 4-ethenyl-4-methyl-3-(1-methylethenyl)-1-(1-methylethyl)-, (3R-trans)- 19.85 C15H24 0.04±0.01a 0.03±0.01a Nd Nd Nd Nd Nd Nd
Copaene 20.82 C15H24 0.24±0.04a 0.27±0.07a 0.57±0.03a 0.57±0.17a 0.68±0.22a 0.85±0.08a 0.52±0.08a 0.47±0.09a
Caryophyllene 21.83 C15H24 0.66±0.12a 0.68±0.16a 1.59±0.05a 1.75±0.58a 2.1±0.73a 2.5±0.16a 1.54±0.32a 1.37±0.22a
Humulene 22.63 C15H24 0.05±0.01a 0.06±0.01a Nd Nd Nd 0.3±0.03 Nd 0.11±0.01
Benzene, 1-(1,5-dimethyl-4-hexenyl)-4-methyl- 23.18 C15H22 0.13±0.02a 0.15±0.03a 0.59±0.05a 0.56±0.16a 0.66±0.26a 0.85±0.11a 0.56±0.1a 0.5±0.12a
1,3-Cyclohexadiene, 5-(1,5-dimethyl-4-hexenyl)-2-methyl-, [S-(R*,S*)]- 23.47 C15H24 0.5±0.14ab 0.46±0.38a 1.49±0.04a 1.69±0.57a 2.03±0.73a 2.88±0.36a 1.6±0.32a 1.9±0.5a
Bicyclo[3.1.1]hept-2-ene, 2,6-dimethyl-6-(4-methyl-3-pentenyl)- 23.47 C15H24 0.5±0.14ab 0.21±0.32a 1.49±0.04a 1.69±0.57a 2.03±0.73a 2.88±0.36a 1.6±0.32a 1.9±0.5a
β-Curcumene 23.47 C15H24 0.5±0.14a 0.72±0.12b 1.49±0.04a 1.69±0.57a 2.03±0.73a 2.88±0.36a 1.6±0.32a 1.9±0.5a
α-Farnesene 12.43 C15H24 0.19±0.03 Nd 0.56±0.03a 0.54±0.15a 0.72±0.26a 0.91±0.1a 0.72±0.13a 0.53±0.14a
β-Bisabolene 23.74 C15H24 0.12±0.03a 0.14±0.03a 0.42±0.02a 0.39±0.11a 0.5±0.19a 0.61±0.07a 0.64±0.17a 0.38±0.1b
1H-3a,7-Methanoazulene, octahydro-3,8,8-trimethyl-6-methylene-, [3R-(3α,3aβ,7β,8aα)]- 24.05 C15H24 0.17±0.04a 0.19±0.03a Nd 0.63±0.18 Nd Nd Nd Nd
Cedrene 24.05 C15H24 0.17±0.04a 0.13±0.1a Nd Nd Nd Nd Nd Nd
β-Myrcene 9.11 C10H16 Nd 0.78±0.7 Nd Nd Nd Nd Nd Nd
Alloaromadendrene 23.74 C15H24 Nd 0.22±0.04 Nd Nd Nd Nd Nd Nd
1,3-Cyclopentadiene, 5-(1-methylethylidene)- 7.6 C8H10 Nd Nd 0.19±0.05a 0.18±0.01a 0.26±0.1 Nd 0.14±0.02 Nd
3-Carene 9.34 C10H16 Nd Nd 0.21±0.01 Nd Nd 0.26±0.03 Nd 0.13±0.02
D-Limonene 12.18 C10H16 Nd Nd 2.68±0.03a 2.78±0.84a 3.71±1.51a 2.49±2.13ab 2.55±0.4 Nd
Cyclohexene, 1-methyl-4-(1-methylethenyl)-, (S)- 12.18 C10H16 Nd Nd 2.68±0.03a 2.78±0.84a Nd 2.49±2.13 2.55±0.4 Nd
Benzene, 1-methyl-4-(1,2,2-trimethylcyclopentyl)-, (R)- 23.2 C15H22 Nd Nd 0.59±0.05a 0.56±0.16a 0.66±0.26a 0.85±0.11a 0.56±0.1a 0.5±0.12a
(E)-β-Famesene 24.06 C15H24 Nd Nd Nd Nd 0.79±0.28a 0.99±0.11a Nd 0.65±0.2
1,3,5-Cycloheptatriene 4.66 C7H8 Nd Nd Nd Nd Nd 0.24±0.02 Nd Nd
Styrene 8.31 C8H8 Nd Nd Nd Nd Nd 0.71±0.07 Nd 0.41±0.07
1,3,5,7-Cyclooctatetraene 8.3 C8H8 Nd Nd Nd Nd Nd 0.71±0.07 Nd Nd
Others (8)
Toluene 4.7 C7H8 0.09±0.01a 0.08±0.02a Nd Nd Nd 0.24±0.02 Nd 0.24±0.04
Ethylbenzene 7.33 C8H10 0.09±0.01a 0.08±0.01a Nd Nd Nd Nd Nd Nd
p-Xylene 7.61 C8H10 0.27±0.05b 0.25±0.06a 0.19±0.05a 0.18±0.01a Nd 0.2±0.05 0.14±0.02a 0.15±0.01a
Benzene, 1,3-dimethyl- 12.11 C8H10 0.27±0.05 Nd Nd 0.18±0.01 Nd Nd Nd Nd
o-Xylene 7.61 C8H10 0.19±0.16a 0.12±0.16a Nd Nd Nd Nd Nd Nd
o-Cymene 12.1 C10H14 0.32±0.05a 0.32±0.1a 0.31±0.01b 0.38±0.05a 0.45±0.19a 0.45±0.05a 0.31±0.04a 0.22±0.02b
Benzene, 1-methyl-3-(1-methylethyl)- 13.11 C10H14 0.32±0.05 Nd Nd Nd Nd Nd Nd Nd
2-Methoxy-5-methylphenol 11.58 C8H10O2 Nd Nd Nd Nd Nd Nd Nd 3.73±0.34
All contents 50.66 50.97 93.05 98.51 75.29 112.41 58.08 68.8
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The results are expressed as the mean ± S.D. (n = 3). Nd indicates no detection. Different lowercase letters indicated significant differences between groups (P<0.05), while the same letter indicated no significant difference (P>0.05).

Biogenic amine analysis

It showed that the contents of various biogenic amines in CC group were significantly lower than those in PC group during the 41 d storage period (P<0.05) and the total bioamine content in CC group was relatively lower than that in PC group at every stages (Table 7). The contents of tryptamine, cadaverine, phenylethylamine, putrescine, histamine and tyramine in CC group were significantly lower than those in PC group (P<0.05), while spermidine was significantly higher than that in PC group (P<0.05) (Fig 1). During storage, collagen casings could reduce the production of biogenic amines compared with pig casings, thus effectively improving the safety of fermented sausages.

Table 7. Changes of biogenic amine contents.

Content, mg/kg Group 11 d 21 d 41 d 61 d
Tryptamine PC 2.0±0.04Aa 3.6±0.08Ba 13.4±0.10Ca 15.7±0.33Da
CC 1.4±0.01Ab 2.1±0.12Bb 2.3±0.06Cb 3.2±0.02Db
Phenethylamine PC 3.1±0.006Aa 4.7±0.008Ba 8.5±0.07Ca 4.8±0.002Da
CC 3.8±0.002Ab 4.7±0.01Ba 7.2±0.003Cb 3.5±0.008Db
Putrescine PC 288.2±0.10Aa 225.8±0.27Ba 281.8±2.41Ca 188.8±0.07Da
CC 77.9±0.02Ab 104.5±0.15Bb 118.5±0.02Cb 185.4±0.31Db
Cadaverine PC 142.9±0.51Aa 139.6±0.16Ba 304.0±2.78Ca 403.1±0.04Da
CC 86.3±0.007Ab 107.5±0.13Bb 106.3±0.16Cb 223.4±0.31Db
Histamine PC 14.5±0.06Aa 21.3±0.04Ba 34.2±0.28Ca 22.9±0.02Da
CC 8.7±0.03Ab 8.2±0.04Bb 12.0±0.02Cb 9.6±0.03Db
Tyramine PC 360.1±0.30Aa 573.1±0.37Ba 600.4±5.33Ca 567.0±0.32Ba
CC 348.1±0.14Ab 305.9±1.51Bb 310.7±1.19Cb 411.5±0.25Db
Spermidine PC 3.7±0.007Aa 3.4±0.01Ba 4.8±0.06Ca 3.5±0.004Ba
CC 4.4±0.04Ab 4.5±0.07Ab 4.1±0.03Bb 6.1±0.01Cb
Total Content, mg/kg PC 814.5 971.5 1246.8 1205.7
CC 530.6 537.4 561.1 842.7
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The data of different bioamines in the table were the mean ± S.D. and the total content were the mean. The three fermented sausage per experimental group of three independent experiments. Different capital letters indicated significant differences over time within the same group (P<0.05) and different lowercase letters indicated significant differences between groups at the same time (P<0.05), while the same letter indicated no significant difference (P>0.05).

Fig 1. Biogenic amine content heatmap analysis for fermented sausages during storage.

Fig 1

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Each column in heat map represented a different point in time. The ordinate was the relative abundance of each group. The closer the color is to red, the higher the relative abundance of corresponding biogenic amines, and the closer the color is to blue, the lower the relative abundance of corresponding biogenic amines.

Discussion

This study focused on the effect of CC on biogenic amine contents and other quality indicators during the fermentation of sausage. Since lactic acid bacteria produce acid in fermentation process, the pH value of fermented sausage decreases. When it reaches the lowest point, the growth of the starter strain is inhibited, the acid production capacity decreases, and the pH value slowly increases [20]. The pH value of CC group was lower than that of PC group at all stages after fermentation. This phenomenon may be caused by the different tissue structures of these two casings, leading to differences in the contents of amines and ammonia produced by proteolysis and lipid metabolism [21]. Compared with PC group, the lower pH value could inhibit the growth of putrefying bacteria in CC group, which was conducive to maintain quality of fermented sausage during storage. We found that the different color values of fermented sausage made by collagen casings were lower than those of pig casings. Collagen casings have lower transparency, larger thickness, lower elasticity, worse air permeability and worse binding ability with oxygen than natural casings, as they are made of extracted collagen and ingredients [22]. Therefore, it is necessary to improve the color of fermented sausage by adjusting the tissue structure of the collagen casings in further studies.

Brandy et al. revealed that the texture was related to mouthfeel of meat products [23], which was an important factor determining whether meat products that can be accepted by consumers. In our study, CC group exhibited improved texture characteristics of fermented sausages compared with PC group. During storage, the decrease of water activity compacted the structure of the sausage and increases its hardness gradually [24]. Meanwhile, the elasticity of PC group was better than that of CC group, because the permeability of collagen casings was stronger [5]. However, the poor elasticity of collagen casing made the sausages more compact and the small molecular products (oligopeptides and small amino acids) were not easily deteriorated or lost after fermentation, which made the sausage have better gumminess [25]. According to the analysis as mentioned above, the structure of fermented sausage in CC group was more delicate and compact than that in PC group.

The main sources of volatile flavor components in fermented sausage are from adding spices, and flavor compounds formed by the degradation of protein, fat and carbohydrate by enzymes [26, 27]. The contents and types of terpenes which from spices were more in each group. The main components of pepper flavor are β-cardene, limonene, β-pinene and β-laurene [28, 29]. All these substances were detected in our experimental groups, which were mainly from the black pepper in processed raw materials. And it contributes greatly to the flavor of fermented sausage. In addition, carbohydrate metabolism had a great influence on the flavor formation of fermented sausage, which was mainly completed by LAB [30]. Compounds derived from carbohydrate metabolism such as ethanol, 3-hydroxy-2-butanone and acetic acid were detected in two experimental groups. Compared with collagen casings, the good air permeability of pig casings encourages microorganisms such as lactobacillus to proliferate and metabolize carbohydrates, forming flavor substances such as 3-hydroxy-2-butanone.

We surprisingly found that collagen casings can effectively reduce the contents of biogenic amines in fermented sausage, thus improving the safety of fermented sausage products. During storage, the total contents of biogenic amines in fermented sausage of CC group were all lower than those in PC group. Among them, the content of phenylethylamine in two groups increased firstly and then decreased during storage time. Intermediate storage appeared rise phenomenon because of high temperature during the early stage of fermentation which resulted in stronger activity of phenylalanine decarboxylase and the stronger phenylalanine decarboxylation caused the more accumulation of phenylethylamine finally [31]. Meanwhile, phenylethylamine no longer accumulated in large quantities and gradually reduced by the oxidases produced by microorganisms when decarboxylase activity declined at low temperature storage period (4°C). Otherwise, putrescine is usually used together with cadaverine as the quality indicator of fermented sausage [32]. We thought that the better air permeability of pig casings made the total aerobic mesophilic bacteria in PC group higher than that in CC group in the early stage and then altered the function of the microbial enzyme system that decomposes protein to produce free amino acids. These free amino acids were precursors of biogenic amines, such as ornithine, which resulted in a high content of putrescine [33, 34], which was the main reason for the difference in the biogenic amine content between the two groups [19].

Conclusions

In this study, two types of casings of fermented sausage were compared and analyzed by the determination of the biogenic amines and quality characteristics. We revealed that collagen casings improved the texture characteristics of fermented sausage, decreased the pH value, and reduced the content of biogenic amines in fermented sausage. However, collagen casing has no special effect on microbial growth and the formation of volatile flavor compounds. In conclusion, this study studied the positive influence of collagen casings on the quality of fermented sausage from the perspective of product safety, and provided a theoretical basis for the application of casings.

Supporting information

S1 Table. Data of color measurement during processing and storage of fermented sausages.

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S2 Table. Data of water activity measurement during processing and storage of fermented sausages.

(XLSX)

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S3 Table. Data of pH during processing and storage of fermented sausages.

(XLSX)

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S4 Table. Data of microbiological indicators during processing and storage of fermented sausages.

(XLSX)

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S5 Table. Data of TPA measurement during storage of fermented sausages.

(XLSX)

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S6 Table. Determination results of volatile flavor substances during storage of fermented sausages.

(XLSX)

Click here for additional data file. (67.8KB, xlsx)
S7 Table. Data of biogenic amine measurement during storage of fermented sausages.

(XLSX)

Click here for additional data file. (10.7KB, xlsx)
S8 Table. Basic data of heatmap.

(XLSX)

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Data Availability

All relevant data are within the paper and its Supporting Information files.

Funding Statement

This work was funded by Food Science and Engineering College Science and Technology Plan Project of Inner Mongolia Agricultural University, China (SPKJ201904) (https://spy.imau.edu.cn/)and Inner Mongolia Natural Science Foundation Program of China (No.2021MS03012)(http://kjt.nmg.gov.cn/). And Yan D received them. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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Decision Letter 0

A M Abd El-Aty

1 Oct 2021

PONE-D-21-28210Effect of Collagen Casing on the Quality Characteristics of Fermented SausagePLOS ONE

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Comments to the Author

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Reviewer #1: Yes

Reviewer #2: Partly

Reviewer #3: Partly

Reviewer #4: Yes

Reviewer #5: Partly

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

Reviewer #4: Yes

Reviewer #5: Yes

**********

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**********

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**********

5. Review Comments to the Author

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Reviewer #1: Plos One

PONE-D-21-28210

Effect of Collagen Casing on the Quality Characteristics of Fermented Sausage

Dear Editor,

The article deals with the effect of casing types on the quality characteristics and biogenic amine contents of the fermented sausages. The topic is interesting. It can be published in your journal after necessary corrections have been done. My specific comments and questions are below.

- What was the main purpose of this research? Why was it thought that casing type would affect the formation of biogenic amines? This idea needs to be clearly stated in the introduction section.

- Some information about biogenic amine and health issues and legal limits should be given in the introduction section.

- Line 55: what do you mean “too high”?

- Line 128: Give the incubation conditions!

- Line 131: “total number of colonies” should be “total aerobic mesophilic bacteria” throughout the manuscript.

- Line 141: Much more information about the determination of biogenic amines including the HPLC conditions should be given in the text.

- Fig 1 and Fig 1b should be given as Tables!

- Line 179-180: Please extent the sentence! It is not clear!

- Table 3: Give the total! And discuss it!

- Why the variation in pH values were important but in LAB not?

- Discussion section should be improved!

Reviewer #2: -In the manuscript PONE-D-21-28210, the authors made a comparative analysis of casing effects on sausage quality focusing on biogenic amines and selected physical characteristics. Overall, the limited contents and experiments made the manuscript very facile. Hence, the authors are recommended to evaluate the variations of other bio-molecules, particularly fats, fatty acid, and other common volatile components as well. These molecules are one of the determinants of sausage quality and hence, analyzing the effects of casing could improve the value of the manuscript. The authors should also specify the type of meat used during sausage production, and explicitly describe the HPLC protocol they have followed during the analysis/quantification of the biogenic amines. In addition, the morphology/texture of the sausage samples at each processing and storage time should be supported by optical microscope (SEM) data.

Reviewer #3: Manuscript (PlosOne)

"Effect of collagen casing on the quality characteristics of fermented sausage"

Hereby my comments to the manuscript:

I think you should add an extra keyword that included the color and texture measurements

General remark for the introduction:

Describe what casings are used now for the production of fermented sausages. You do write that there are different types of edible casings, but it would be clearer for the story if you mention that the pig casing is used as a reference, since fermented sausages are made with natural casings.

Line 55: an individual’s intake is too high? -> Do you have numbers? What is too high?

Line 64: I do not agree with the sentence that collagen casings has a similar structure to natural casing! Maybe the collagen type is similar, but how the structures and the collagen network are constructed differ quite a bit. In addition, natural intestine also contains other proteins, such as elastin, while these are not present in the collagen casings. And it is precisely the presence of these other proteins that gives the natural intestine a unique structure and texture characteristics.

Line 79: involved in this study should be replaced by investigated

Line 96: Which type/ brand/ diameter of the collagen casing did you use? Furthermore, with what equipment did you stuff the casings with meat dough? What was the filling pressure, etc? Was this kept the same for the two casing types?

Line 109: How many samples did you measure for the color analysis? What apparatus was used for shredding? Why shredding the samples for the colour analysis. Why didn’t you cut a slice of the sausage and then measure the surface colour?

Line 118: this vessel is specific for the water activity meter?

Line 123: I would at the pH of the saline solution and the concentration/ molarity of the salt solution.

Texture profile analysis: what were the settings of the texture analyser: for example speed, compression % load cell, etc. I think it is good to mention that the gumminess and chewiness of the samples are not measured, but are calculated values.

Line 167: Why is there no results description regarding the measured L, a, b values.

Line 187 – 190: Is a repetition. Already described in material and methods.

Line 213 – 214: Here you already provide an explanation for the results obtained and a conclusion while you do not do that with the other results

Line 234: Do you mean difference in permeability?

Line 240: replace which are to: as they are made

Line 241 -242: How? Are you going to make your own prefabricated collagen casing? Production of a prefabricated collagen casing is a proprietary/ specific process.

Line 243: Brandy et al. -> Brady et al. Furthermore “believed” does not sound very scientific.

Line 245 – 246: With this statement you say that PC is your reference -> is that right? Otherwise you can’t make the statement.

Line 248 – 251: what does the elasticity of the meat stuffed in PC have to do with the permeability of the casing? You are talking about elasticity of the casing, but you did not measure this?

Line 250: collagen casing is not an artificial casing -> artificial casing is a polyamide casing.

Line 251 – 253: Why? And are these small products broken down at PC?

Line 260: permeability for what? Oxygen/ water/ etc.

In general: I would rewrite the discussion part of texture. Now very confusing and not clear.

Reviewer #4: In this study, the effects of collagen casings and natural casings (pig casings) on the biogenic amine content and other quality characteristics of fermented sausage during storage were analyzed. Compared with natural casings, collagen casings could improve the quality characteristics of fermented sausage and reduce the content of biogenic amines in fermented sausage, The study is novel and the results are valuable for the choice of casings in industrial production.

But there are several problems that need to be addressed.

1. How many samples used to measure meat quality traits?

2. In the statistical analysis, “including all interactions between processing and time”, how is that present in the data?

3. There were significant differences in the same group at different periods. So, need to add some discussion about the effect of storage period on biogenic amine content and other quality characteristics of fermented sausage.

4. How collagen casings could reduce the content of biogenic amines in fermented sausage?

5. In the discussion, more theories and references are needed to support the conclusion.

Reviewer #5: The research is important in terms of knowledge about the differences between the sausages in various casings. However, the manuscript cannot be published in a current state. The Discussion is poor and is worth deeper investigation. The conclusion concerning improved texture (harder is better?) is questionable. The statement that CC influenced lower biogenic amines production should be discussed regarding increased levels of BA in PC sausages at the beginning of the fermentation, which means that casings might have contained BA and they were only transferred into the sausage. Taking into account that this is the main conclusion, the manuscript is hard to accept.

Below are my forther detailed comments:

Line 51 why is thermal stability of biogenic amines important if the fermented sausages are usually produced and eaten raw? Do you have in mind any kind of fermented sausages intended to fry/cook?

Line 73 the reference number is missing at Skaliac et al

Line 74 US-EPA PAHs should be explained

Line 75 P<0.05 P should be in italics

Line 76 It should be Zajac et al not Marzena and the number should be at the names as in like 73

Line 125 the space is missing between pH and 4.0

Line 128 The microbiological analysis should be properly described: type of diluting solution, names of microbiological media used, conditions of incubations (time/temperature)

Line 135 the type of texturometer should be mentioned

Line 138 1 cm×1 cm3 3 is missing at first cm

Line 139 the compression conditions should be included

Line 141. The type of chromatograph and detector used should be included. The methods could be described in more detail

Table 1 the significance letters should be at mean values not at errors

Line 170 The information what PC or CC stand for should be added – the table should be self-explanatory

Line 183-190 this information was already given in Materials and methods section

Line 202 There were significant differences between different capital letters in the same group at different periods (P<0.05). There were

significant differences in different lowercase letters among different groups in the

same period (P<0.05). – it should be shortened eg. Capital letters indicate….

Line 234 There were several studies in which fermented sausages produced in collagen casings were produced. Was the same low pH in CC detected? It would be worth discussing

Line 240 This comment would be adequate if the casings production would be a subject of the study. In my opinion it is not suitable here.

Line 243 It should be paraphrased. Tenderness is one of the texture parameters – there is nothing to believe in

Line 245 Why was it improved? Is harder better?

**********

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Reviewer #1: No

Reviewer #2: No

Reviewer #3: No

Reviewer #4: No

Reviewer #5: No

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PLoS One. 2022 Feb 3;17(2):e0263389. doi: 10.1371/journal.pone.0263389.r002

Author response to Decision Letter 0

10 Nov 2021

Dear editor:

Thank you for providing these comments on our submission entitled “Effect of Collagen Casing on the Quality Characteristics of Fermented Sausage”. We appreciate the constructive comments from you and five reviewers. We have studied the comments carefully and provided a detailed response to each point raised.

Response to Editor

Question 1: Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming.

Response to question 1: Thank you for your suggestion. We have revised our manuscript followed the style requirement of the journal .

Question 2: We suggest you thoroughly copyedit your manuscript for language usage, spelling, and grammar. If you do not know anyone who can help you do this, you may wish to consider employing a professional scientific editing service.

Response to question 2: Thanks for your helpful suggestions. We sought for the help from AJE (American Journal Experts) to refine our paper, the order number is D3C61K32.

Question 3: We understand that you purchased meat, extracts from local markets for this study. In your Methods section, please provide additional regarding the source of this material. Please provide the geographic coordinates and names of the purchase locations (e.g., stores, markets), if available, as well as any further details about the purchased items (e.g., lot number, source origin, description of appearance) to ensure reproducibility of the analyses.

Response to question 3: Thank you for your careful review. We have added relevant content in our revised manuscript (lines 86-91, 97-101, page 5).

Question 4: We note that you have provided funding information that is not currently declared in your Funding Statement. However, funding information should not appear in the Acknowledgments section or other areas of your manuscript. We will only publish funding information present in the Funding Statement section of the online submission form. Please remove any funding-related text from the manuscript and let us know how you would like to update your Funding Statement.

Response to question 4: Thank you for your suggestion. We were apologized for this error message, we have deleted this part from our manuscript and either from the submission system.

Question 5: Upon re-submitting your revised manuscript, please upload your study’s minimal underlying data set as either Supporting Information files or to a stable, public repository and include the relevant URLs, DOIs, or accession numbers within your revised cover letter.

Response to question 5: All data are in our manuscript.

Question 6: PLOS requires an ORCID iD for the corresponding author in Editorial Manager on papers submitted after December 6th, 2016. Please ensure that you have an ORCID iD and that it is validated in Editorial Manager.

Response to question 6: Thanks for your suggestion. We have confirmed and uploaded the information on the system.

Response to Reviewer 1

Question 1: What was the main purpose of this research? Why was it thought that casing type would affect the formation of biogenic amines? This idea needs to be clearly stated in the introduction section.

Response to question 1: Thanks for your suggestion. We have added relevant content in our revised manuscript (lines 91-94, 83-86, page 5).

Question 2: Some information about biogenic amine and health issues and legal limits should be given in the introduction section.

Response to question 2: We searched relevant domestic and foreign laws and regulations. At present, there are no laws and regulations on the content limit standard of other biological amines except histamine. And most of the laws and regulations on histamine focus on fish and fish products. Such as “Zhao JIadi. Screening of bioamine-degrading bacteria and its application in soybean sauce [D]. Jiangnan University, 2020.”

Question 3: Line 55: what do you mean “too high”?

Response to question 3: Thank you for your careful review. We have revised this sentence (lines 64-65, page 4).

Question 4: Line 128: Give the incubation conditions.

Response to question 4: Thanks for your suggestion. We have added relevant content in our revised manuscript (lines 157-158, page 8).

Question 5: Line 131: “total number of colonies” should be “total aerobic mesophilic bacteria” throughout the manuscript.

Response to question 5: Sorry for this mistake. We have corrected the mistake in our revised manuscript (line 161, 223, 315, page 8, 12, 18).

Question 6: Much more information about the determination of biogenic amines including the HPLC conditions should be given in the text.

Response to question 6: Thank you for your constructive suggestion. We have added relevant content in our revised manuscript (lines 180-187, pages 9-10; Table 1).

Question 7: Fig 1 and Fig 1b should be given as Tables!

Response to question 7: Thank you for your constructive suggestion. We have revised these parts in Table.2 and Table.3 of our revised manuscript.

Question 8: Line 179-180: Please extent the sentence! It is not clear!

Response to question 8: Thank you for your careful review. We have modified this sentence of our revised manuscript (lines 223-226, page 12).

Question 9: Table 3: Give the total! And discuss it.

Response to question 9: Thank you for your suggestion. We have added relevant content in our revised manuscript (lines 256-258, 304-305, pages 15-16,18-19).

Question 10: Why the variation in pH values were important but in LAB not?

Response to question 10: Thank you for your careful review. We have revised some contents in our revision (lines 281-283, page 17-18).

Question 11: Discussion section should be improved!

Response to question 11: Thanks for your suggestion. We have improved and added many contents in our revision (Discussion Part).

Response to Reviewer 2

The authors made a comparative analysis of casing effects on sausage quality focusing on biogenic amines and selected physical characteristics. Overall, the limited contents and experiments made the manuscript very facile.

Question 1: The authors are recommended to evaluate the variations of other bio-molecules, particularly fats, fatty acid, and other common volatile components as well. These molecules are one of the determinants of sausage quality and hence, analyzing the effects of casing could improve the value of the manuscript.

Response to question 1: Thanks for your constructive suggestion. We have conducted most of the indexes which you mentioned in our next study to further explore the effect of collagen casings on fat and protein decomposition and flavor of fermented sausages. And the relative article named "The Effect of Collagen Casing on the Flavor Formation of Fermented Sausage" is under modified and prepared to submitted “Meat Science”. We hope you could pay attention to this work if you were interested in our research field.

Question 2: The authors should also specify the type of meat used during sausage production, and explicitly describe the HPLC protocol they have followed during the analysis/quantification of the biogenic amines.

Response to question 2: Thanks for your suggestion. We have added relevant content in our revised manuscript (lines 180-187, pages 9-10; Table 1).

Question 3: The morphology/texture of the sausage samples at each processing and storage time should be supported by optical microscope (SEM) data.

Response to question 3: Thank you for your suggestion. Our study focused on the effect of casings on the quality characteristics of fermented sausage mainly. We will use the optical microscope to conduct in-depth research on the tissue structure of casing in the subsequent experiments.

Response to Reviewer 3

"Effect of collagen casing on the quality characteristics of fermented sausage"

Hereby my comments to the manuscript:

Question 1: I think you should add an extra keyword that included the color and texture measurements.

Response to question 1: Thanks for your suggestion. We have added relevant content in our revised manuscript (line 46, page 3).

Question 2: Describe what casings are used now for the production of fermented sausages. You do write that there are different types of edible casings, but it would be clearer for the story if you mention that the pig casing is used as a reference, since fermented sausages are made with natural casings.

Response to question 2: Thanks for your suggestion. We have revised the paragraph (lines 72-73, page 4).

Question 3: Line 55: an individual’s intake is too high? -> Do you have numbers? What is too high?

Response to question 3: Thank you for your careful review. We have revised this inappropriate statement (lines 64-65, page 4).

Question 4: Line 64: I do not agree with the sentence that collagen casings has a similar structure to natural casing! Maybe the collagen type is similar, but how the structures and the collagen network are constructed differ quite a bit. In addition, natural intestine also contains other proteins, such as elastin, while these are not present in the collagen casings. And it is precisely the presence of these other proteins that gives the natural intestine a unique structure and texture characteristics.

Response to question 4: Sorry for this mistake. We have corrected the mistake in our revised manuscript. (lines 75-76, page 5).

Question 5: Line 79: involved in this study should be replaced by investigated.

Response to question 5: Thank you for your suggestion. We have revised this mistake (line 92, page 5).

Question 6: Line 96: Which type/ brand/ diameter of the collagen casing did you use? Furthermore, with what equipment did you stuff the casings with meat dough? What was the filling pressure, etc? Was this kept the same for the two casing types? Line 96: Which type/ brand/ diameter of the collagen casing did you use? Furthermore, with what equipment did you stuff the casings with meat dough? What was the filling pressure, etc? Was this kept the same for the two casing types?

Response to question 6: Thank you for your careful review. We have added relevant content in our revised manuscript (lines 112-116, pages 6).

Question 7: Line 109: How many samples did you measure for the color analysis? What apparatus was used for shredding? Why shredding the samples for the color analysis. Why didn’t you cut a slice of the sausage and then measure the surface color?

Response to question 7: Sorry for this mistake. We have corrected the mistake in our revised manuscript (lines 134-137, page 7).

Question 8: Line 118: this vessel is specific for the water activity meter?

Response to question 8: Thank you for your careful review. The water activity meter has special vessel. We have added some contents in our manuscript (lines 147-148, page 8).

Question 9: Line 123: I would at the pH of the saline solution and the concentration/ molarity of the salt solution. Texture profile analysis: what were the settings of the texture analyzer: for example speed, compression % load cell, etc. I think it is good to mention that the gumminess and chewiness of the samples are not measured, but are calculated values.

Response to question 9: Thank you for your careful review. We have added some contents in our revision (lines 151-152, 169-172, pages 8, 9). According to the actual situation, the viscosity and chewiness data of the texture tester were obtained directly after the test without calculation.

Question 10: Line 167: Why is there no results description regarding the measured L, a, b values.

Response to question 10: Thanks for your suggestion. We have modified this part (lines 209-213, pages 11).

Question 11: Line 187-190: Is a repetition. Already described in material and methods.

Response to question 11: Thanks for your careful review. We have deleted this part in our revision.

Question 12: Line 213-214: Here you already provide an explanation for the results obtained and a conclusion while you do not do that with the other results

Response to question 12: Thank you for your careful review. We have revised this sentence in our manuscript (lines 293-301, page 17).

Question 13: Line 234: Do you mean difference in permeability?

Response to question 13: Thank you for your careful review. The structure of different casings makes casings different in air permeability.

Question 14: Line 240: replace which are to: as they are made.

Response to question 14: Thanks for your suggestion. we have corrected this part in our revised manuscript (lines 286-287, page 17).

Question 15: Line 241 -242: How? Are you going to make your own prefabricated collagen casing? Production of a prefabricated collagen casing is a proprietary/ specific process.

Response to question 15: Thank you for your constructive suggestion. We think there are many ways to improve the color of fermented sausage. Such as, we could improve the structure of collagen casing and also could modify the formula of fermented sausage.

Question 16: Line 243: Brandy et al. -> Brady et al. Furthermore “believed” does not sound very scientific.

Response to question 16: Thank you for your careful review. We have corrected this part in our revised manuscript (line 290, page 17).

Question 17: Line 245 – 246: With this statement you say that PC is your reference -> is that right? Otherwise you can’t make the statement.

Response to question 17: Thanks for your suggestion. The main purpose of our research was to study the characteristics of collagen casings and pig casings on fermented sausage quality and pig casings were used as the control group.

Question 18: Line 248 – 251: what does the elasticity of the meat stuffed in PC have to do with the permeability of the casing? You are talking about elasticity of the casing, but you did not measure this?

Response to question 18: Thank you for your careful review. We have corrected this part in Table.5 of our revised manuscript.

Question 19: Line 250: collagen casing is not an artificial casing -> artificial casing is a polyamide casing.

Response to question 19: Thank you for your suggestion. We searched relevant literature, which pointed out that collagen casings are artificial casings. Patricia Suurs, Shai Barbut. Collagen use for co-extruded sausage casings – A review. Trends in Food Science & Technology. 2020; 102: 91-101. doi: 10.1016/j.tifs.2020.06.011

Question 20: Line 251 – 253: Why? And are these small products broken down at PC?

Response to question 20: Thanks for your review. We have added some content in our revised manuscript. According to the following discussion on the content of biogenic amine in fermented sausage could also explain your problem (lines 296-299, 316-319, page 17,18).

Question 21: Line 260: permeability for what? Oxygen/ water/ etc.

Response to question 21: Thank you for your careful review. We have corrected this part in our revised manuscript (lines 285-286, 315, pages 17,18).

Question 22: In general: I would rewrite the discussion part of texture. Now very confusing and not clear.

Response to question 22: Thanks for your suggestion. We have revised the paragraph (lines 293-301, page 17).

Response to Reviewer 4

In this study, the effects of collagen casings and natural casings (pig casings) on the biogenic amine content and other quality characteristics of fermented sausage during storage were analyzed. Compared with natural casings, collagen casings could improve the quality characteristics of fermented sausage and reduce the content of biogenic amines in fermented sausage, The study is novel and the results are valuable for the choice of casings in industrial production.

Question 1: How many samples used to measure meat quality traits?

Response to question 1: Thank you for your careful review. We have added some content in our revised manuscript (lines 127-130, page 7; Table 2, Table 3, Table 4, Table 5, Table 6).

Question 2: In the statistical analysis, “including all interactions between processing and time”, how is that present in the data?

Response to question 2: Thanks for your suggestion. We have revised this paragraph (lines 192-195, page 10).

Question 3: There were significant differences in the same group at different periods. So, need to add some discussion about the effect of storage period on biogenic amine content and other quality characteristics of fermented sausage.

Response to question 3: Thank you for your suggestion. This research focused on the quality characteristics of fermented sausage in different casings. The reason why there is no detailed analysis of the quality of fermented sausage in different periods is to avoid the separation of our main purpose.

Question 4: How collagen casings could reduce the content of biogenic amines in fermented sausage?

Response to question 4: Thank you for your careful review. There are not many relevant literatures to reveal the effect of collagen casings on fermented sausage bioamines. The starting point for our research was to explore whether collagen casing has an effect on the bioamine production of fermented sausage. The results showed that collagen casings can effectively reduce biogenic amine production, and this phenomenon was discussed and analyzed in discussion part (lines 304-313, page 17-18).

Question 5: In the discussion, more theories and references are needed to support the conclusion.

Response to question 5: Thanks for your suggestion. We have improved and added many contents in our revision (Discussion Part).

Response to Reviewer 5

The research is important in terms of knowledge about the differences between the sausages in various casings. However, the manuscript cannot be published in a current state. The Discussion is poor and is worth deeper investigation. The conclusion concerning improved texture (harder is better?) is questionable. The statement that CC influenced lower biogenic amines production should be discussed regarding increased levels of BA in PC sausages at the beginning of the fermentation, which means that casings might have contained BA and they were only transferred into the sausage.

Question 1: Line 51 why is thermal stability of biogenic amines important if the fermented sausages are usually produced and eaten raw? Do you have in mind any kind of fermented sausages intended to fry/cook?

Response to question 1: Thanks for your suggestion. The fermented sausage made in this study was mainly used to eat raw. Therefore, the effect of cooking and frying on biogenic amine formation of the fermented sausage was out of the point of this work. However, your suggestion is another good point, we will take consideration in our further experimental study. We hope you could pay attention to our follow-up researches continuously.

Question 2: Line 73 the reference number is missing at Skaliac et al.

Response to question 2: Sorry for this funny mistake. We have corrected the mistake in our revised manuscript (line 89, page 5).

Question 3: Line 74 US-EPA PAHs should be explained.

Response to question 3: Thank you for your careful review. We have added relevant content in our revised manuscript (lines 87-88, page 5).

Question 4: Line 75 P<0.05 P should be in italics.

Response to question 4: Sorry for this mistake. We have deleted this part.

Question 5: Line 76 It should be Zajac et al not Marzena and the number should be at the names as in like 73.

Response to question 5: Sorry for this mistake. We have corrected the mistake in our revised manuscript (line 89, page 5).

Question 6: Line 125 the space is missing between pH and 4.0

Response to question 6: Sorry for this mistake and thanks for your careful review. We have corrected the mistake in our revised manuscript (line 153, page 8).

Question 7: Line 128 The microbiological analysis should be properly described: type of diluting solution, names of microbiological media used, conditions of incubations (time/temperature).

Response to question 7: Thank you for your careful review. We have revised some contents in our revision (lines 157-166, pages 8-9).

Question 8: Line 135 the type of texture meter should be mentioned.

Response to question 8: Thank you for your careful review. We have revised some contents in our revision (lines 169-170, page 9).

Question 9: Line 138 1 cm×1 cm3 3 is missing at first cm.

Response to question 9: Sorry for this mistake. We have corrected the mistake in our revised manuscript (line 173, page 9).

Question 10: Line 139 the compression conditions should be included.

Response to question 10: Thank you for your careful review. We have revised some contents in our revision (lines 170-172, page 9).

Question 11: Line 141. The type of chromatograph and detector used should be included. The methods could be described in more detail.

Response to question 11: Thanks for your suggestion. We have added some contents in our revision (lines 180-187, pages 9-10).

Question 12: Table 1 the significance letters should be at mean values not at errors.

Response to question 12: Thank you for your careful review. We think adding standard errors makes the data more reasonable for our results.

Question 13: Line 170 The information what PC or CC stand for should be added – the table should be self-explanatory.

Response to question 13: Thank you for your careful review. CC group and PC group respectively represented collagen casings and pig casings group, which we have mentioned at materials and methods part and we have also added this content in our revision as your suggestion. (lines 219, page 12).

Question 14: Line 183-190 this information was already given in Materials and methods section.

Response to question 14: Thank you for your careful review. We have deleted this part in our revision.

Question 15: Line 202 There were significant differences between different capital letters in the same group at different periods (P<0.05). There were significant differences in different lowercase letters among different groups in the same period (P<0.05). – it should be shortened eg. Capital letters indicate….

Response to question 15: Thank you for your careful review. We have revised these in Table.1, Table.2, Table.3, Table.4, Table.5, Table.6, of our revised manuscript.

Question 16: Line 234 There were several studies in which fermented sausages produced in collagen casings were produced. Was the same low pH in CC detected? It would be worth discussing.

Response to question 16: Thanks for your suggestion. We have added some contents in our revision (lines 281-283, page 17-18).

Question 17: Line 240 This comment would be adequate if the casings production would be a subject of the study. In my opinion it is not suitable here.

Response to question 17: Thank you for your careful review. We have revised this part in our revised manuscript (lines 327-329, page 18-19).

Question 18: Line 243 It should be paraphrased. Tenderness is one of the texture parameters – there is nothing to believe in.

Response to question 18: Thank you for your careful review. We have corrected this part in our revised manuscript (line 290, page 17).

Question 19: Line 245 Why was it improved? Is harder better?

Response to question 19: Thanks for your suggestion. We apologized for our written organization skills. The hardness of fermented sausage in CC group was larger than that in PC group, which made fermented sausage more compact. Mei Qin Feng, Jie Zhang & Jian Sun. Effects of imitated Staphylococcus inoculation on quality and oxidation stability of fermented sausage. Food Science, 1-13. doi:10.7506/ SPKX1002-6630-20210805-062. (in Chinese).

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Decision Letter 1

A M Abd El-Aty

1 Dec 2021

PONE-D-21-28210R1Effect of Collagen Casing on the Quality Characteristics of Fermented SausagePLOS ONE

Dear Dr. Duan,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please submit your revised manuscript by Jan 15 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

  • A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

  • A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

  • An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

A. M. Abd El-Aty

Academic Editor

PLOS ONE

AE Comments:

Still, reviewer # 3 is raising some concern over the revised form of the MS. Do go through the comments and amend the MS accordingly. Further,<o:p></o:p>

1- The P letter for statistical value should be an uppercase-italic face letter. Amend throughout the text<o:p></o:p>

2- Line 33: The results showed that with storage time increasing...amend to The results showed that with increasing the storage time, <o:p></o:p>

3- Lines 81-91:Raise a question about the novel aspect of the current study. If there were several studies (2,5, 13, 14, 15), then "what does the current MS add to the existing knowledge"? This Must be discussed and interpreted in the introduction. <o:p></o:p>

4- Full vendor details should include company, city (state) and country. Amend throughout the text<o:p></o:p>

5- Use mL, uL, L, dL throughout the MS<o:p></o:p>

6- Check spaces and punctuation throughout the MS<o:p></o:p>

7- Section: Determination of biogenic amines: You should provide the details of method performance. This should include calibration curves, linearity, LOD, LOQ, accuracy and precision of quantified biogenic amine <o:p></o:p>

8- Data in Tables should be presented as mean +/-SD, not SE<o:p></o:p>

9- Conclusion: Rewrite it again as it repeats what has been stated in the abstract<o:p></o:p>

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #3: All comments have been addressed

Reviewer #5: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #3: Partly

Reviewer #5: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #3: Yes

Reviewer #5: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #3: Yes

Reviewer #5: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #3: Yes

Reviewer #5: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Dear Editor,

The authors have revised their manuscript according to the reviewers' suggestions. Therefore, it can be accepted in its current form.

Reviewer #3: I think the sample size is too limited, especially for the texture analysis. Also the speed of measuring with TPA is rather low.

The story would have been much stronger when microscopy analysis and permeability of the casings was done.

Reviewer #5: I have only one comment regarding the indication of statistical significance. WHat I mean was that the letters should be at mean values and not at errors. e.g. 20a +/- 0.1 and not 20 +/- 0.1a . Using standard errors was correct of course.

**********

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Reviewer #1: No

Reviewer #3: No

Reviewer #5: No

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PLoS One. 2022 Feb 3;17(2):e0263389. doi: 10.1371/journal.pone.0263389.r004

Author response to Decision Letter 1

3 Dec 2021

Dear editor:

Thank you for providing these comments on our submission entitled “Effect of Collagen Casing on the Quality Characteristics of Fermented Sausage”. We appreciate the constructive comments from you and five reviewers. We have studied the comments carefully and provided a detailed response to each point raised.

Response to Reviewer

Question 1: The P letter for statistical value should be an uppercase-italic face letter. Amend throughout the text.

Response to question 1: Thanks for your careful review. We have corrected the mistake in our revised manuscript (line 35, 36, 38, page 2).

Question 2: Line 33: The results showed that with storage time increasing...amend to The results showed that with increasing the storage time.

Response to question 2: Thanks for your suggestion. We have corrected relevant content in our revised manuscript (lines 31, page 2).

Question 3: Lines 81-91: Raise a question about the novel aspect of the current study. If there were several studies (2,5, 13, 14, 15), then "what does the current MS add to the existing knowledge"? This Must be discussed and interpreted in the introduction.

Response to question 3: Thanks for your suggestion. We have added relevant content in our revised manuscript (lines 95-96, pages 5-6).

Question 4: Full vendor details should include company, city (state) and country. Amend throughout the text.

Response to question 4: Sorry for this mistake. We have corrected the mistake in our revised manuscript (line 105-108, 118, 120, 123, 137, 148-149, 155, 161,163, 165,182 pages 6-9).

Question 5: Use mL, uL, L, dL throughout the MS.

Response to question 5: Sorry for this mistake. We have corrected the mistake in our revised manuscript (line 150 page 8).

Question 6: Check spaces and punctuation throughout the MS.

Response to question 6: Thank you for your constructive suggestion. We have added relevant content in our revised manuscript (lines 105, 159, pages 6, 8).

Question 7: Section: Determination of biogenic amines: You should provide the details of method performance. This should include calibration curves, linearity, LOD, LOQ, accuracy and precision of quantified biogenic amine.

Response to question 7: Thank you for your constructive suggestion. We have added relevant content in our revised manuscript (lines 191-195, page 10).

Question 8: Data in Tables should be presented as mean +/-SD, not SE.

Response to question 8: Sorry for this funny mistake. Actually we have presented our data as mean±SD. We have modified mean±SE to mean±SD in our revised manuscript (lines 209, 224, 237, 243, 257, 274 pages 11-14, 16). If you have any doubts, please check our original data uploaded before.

Question 9: Conclusion: Rewrite it again as it repeats what has been stated in the abstract.

Response to question 9: Thank you for your suggestion. We have added relevant content in our revised manuscript (lines 335-337, page 19).

Attachment

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Decision Letter 2

A M Abd El-Aty

9 Dec 2021

PONE-D-21-28210R2

Effect of Collagen Casing on the Quality Characteristics of Fermented Sausage

PLOS ONE

Dear Dr. Duan,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

==============================

ACADEMIC EDITOR: So many files were uploaded. Upload only the last revised MS with response to the Editor's/reviewers comments. All corrections should be highlighted yellow (not in track-changes mode).

==============================

Please submit your revised manuscript by Jan 22 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

  • A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

  • A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

  • An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

A. M. Abd El-Aty

Academic Editor

PLOS ONE

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Reviewers' comments:

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

PLoS One. 2022 Feb 3;17(2):e0263389. doi: 10.1371/journal.pone.0263389.r006

Author response to Decision Letter 2

10 Dec 2021

Dear editor:

Dear editor:

Thank you for providing these comments on our submission entitled “Effect of Collagen Casing on the Quality Characteristics of Fermented Sausage”. We appreciate the constructive comments from you and five reviewers. We have studied the comments carefully and provided a detailed response to each point raised.

Response to Reviewer

Question 1: The P letter for statistical value should be an uppercase-italic face letter. Amend throughout the text.

Response to question 1: Thanks for your careful review. We have corrected the mistake in our revised manuscript (line 35, 36, 38, page 2).

Question 2: Line 33: The results showed that with storage time increasing...amend to The results showed that with increasing the storage time.

Response to question 2: Thanks for your suggestion. We have corrected relevant content in our revised manuscript (lines 30, 31, page 2).

Question 3: Lines 81-91: Raise a question about the novel aspect of the current study. If there were several studies (2,5, 13, 14, 15), then "what does the current MS add to the existing knowledge"? This Must be discussed and interpreted in the introduction.

Response to question 3: Thanks for your suggestion. We have added relevant content in our revised manuscript (lines 94-96, pages 5-6).

Question 4: Full vendor details should include company, city (state) and country. Amend throughout the text.

Response to question 4: Sorry for this mistake. We have corrected the mistake in our revised manuscript (line 104-107, 117, 119, 122, 136, 147-148, 154, 160, 162, 164,181 pages 6-9).

Question 5: Use mL, uL, L, dL throughout the MS.

Response to question 5: Sorry for this mistake. We have corrected the mistake in our revised manuscript (line 149, 158 page 8).

Question 6: Check spaces and punctuation throughout the MS.

Response to question 6: Thank you for your constructive suggestion. We have added relevant content in our revised manuscript (lines 105, 159, pages 6, 8).

Question 7: Section: Determination of biogenic amines: You should provide the details of method performance. This should include calibration curves, linearity, LOD, LOQ, accuracy and precision of quantified biogenic amine.

Response to question 7: Thank you for your constructive suggestion. We have added relevant content in our revised manuscript (lines 190-194, page 10).

Question 8: Data in Tables should be presented as mean +/-SD, not SE.

Response to question 8: Sorry for this funny mistake. Actually we have presented our data as mean±SD. We have modified mean±SE to mean±SD in our revised manuscript (lines 208, 235, 248, 254, 280, 297 pages 11-14, 16). If you have any doubts, please check our original data uploaded before.

Question 9: Conclusion: Rewrite it again as it repeats what has been stated in the abstract.

Response to question 9: Thank you for your suggestion. We have added relevant content in our revised manuscript (lines 358-360, page 19).

Decision Letter 3

A M Abd El-Aty

17 Dec 2021

PONE-D-21-28210R3Effect of Collagen Casing on the Quality Characteristics of Fermented SausagePLOS ONE

Dear Dr. Duan,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

==============================

ACADEMIC EDITOR: The authors have to provide the additional data requested by the diligent reviewer. This will be the last option for revision. Without doing this, the MS will not considered for publication in PLOS ONE.

==============================

Please submit your revised manuscript by Jan 31 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

  • A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

  • A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

  • An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

A. M. Abd El-Aty

Academic Editor

PLOS ONE

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #2: (No Response)

Reviewer #3: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #2: Partly

Reviewer #3: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #2: Yes

Reviewer #3: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #2: No

Reviewer #3: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #2: Yes

Reviewer #3: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #2: Although the authors improved the quality of the manuscript, they did not incorporate the additional data recommended during the past reviews and the manuscript is still superficial. The authors were advised to add data related to the variations of fats, fatty acid, and other common volatile components, but failed to do so claiming they will consider such results for another publication. Besides, results from optical microscope (SEM) analysis are still lacking. With a focus on some biogenic amines and a few physical characteristics only, the manuscript is not in the required standard to be considered for publication.

Reviewer #3: The authors have incorporated the comments. I understand that running extra texture measurements and incorporate permeability measurements is not possible. Maybe a suggestion for future articles. This article is now ready for publishing.

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #2: No

Reviewer #3: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

PLoS One. 2022 Feb 3;17(2):e0263389. doi: 10.1371/journal.pone.0263389.r008

Author response to Decision Letter 3

31 Dec 2021

Dear editor:

Thank you for providing these comments on our submission entitled “Effect of Collagen Casing on the Quality Characteristics of Fermented Sausage”. We appreciate the constructive comments from you and five reviewers. We have studied the comments carefully and provided a detailed response to each point raised.

Response to Reviewer 2

Question 1: The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Response to question 1: We have checked our experiment design and original data, meanwhile have added some experiments on volatile components to make our results experiments more complete.

Question 2: Although the authors improved the quality of the manuscript, they did not incorporate the additional data recommended during the past reviews and the manuscript is still superficial. The authors were advised to add data related to the variations of fats, fatty acid, and other common volatile components, but failed to do so claiming they will consider such results for another publication. Besides, results from optical microscope (SEM) analysis are still lacking. With a focus on some biogenic amines and a few physical characteristics only, the manuscript is not in the required standard to be considered for publication.

Response to question 2: Thank you for your suggestion. we have measured the fermented sausage flavor and showed relevant contents in our revised manuscript (line 36-37, page 2, line 180-184, page 9, 269-286, 15-18, 336-349, page 21-22 and table 6). Our research mainly focused on the effect of different casing types on the quality characteristics of fermented sausage. The research on the organizational structure of casings itself was not the focus of this research. Your valuable suggestions have been involved in our subsequent research. Please keep an eye on our research progress.

Response to Reviewer 3

Question 1: The authors have incorporated the comments. I understand that running extra texture measurements and incorporate permeability measurements is not possible. Maybe a suggestion for future articles. This article is now ready for publishing.

Response to question 1: Thank you very much for your valuable suggestions. We will adopt the measurement of relevant indicators you proposed in the following research. Please keep paying attention to us.

Attachment

Submitted filename: Response to Reviewers.docx

Click here for additional data file. (15.8KB, docx)

Decision Letter 4

A M Abd El-Aty

5 Jan 2022

PONE-D-21-28210R4Effect of Collagen Casing on the Quality Characteristics of Fermented SausagePLOS ONE

Dear Dr. Duan,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

==============================

ACADEMIC EDITOR: Further justifications are needed as raised by the diligent reviewer. Do go through the comments and amend the MS accordingly. Proofread the text for grammar and syntax errors, if any.

==============================

Please submit your revised manuscript by Feb 19 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

  • A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

  • A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

  • An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

A. M. Abd El-Aty

Academic Editor

PLOS ONE

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #2: (No Response)

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #2: Partly

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #2: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #2: No

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #2: With the addition of data related to volatile components, the content of the manuscript has now improved to some extent. Related to this, the authors should clarify the method regarding the analysis of volatile components (lines 180-184). Specifically, the modified method, the identification procedure, and the quantification techniques along with key GC-MS parameters should be described. More importantly, it would be great if the authors incorporate the effect of storage time (21 d, 41 d and 61d) on the variations of these volatile molecules (as they did for biogenic amines).

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #2: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

PLoS One. 2022 Feb 3;17(2):e0263389. doi: 10.1371/journal.pone.0263389.r010

Author response to Decision Letter 4

12 Jan 2022

Dear editor:

Thank you for providing these comments on our submission entitled “Effect of Collagen Casing on the Quality Characteristics of Fermented Sausage”. We appreciate the constructive comments from you and five reviewers. We have studied the comments carefully and provided a detailed response to each point raised.

Response to Reviewer 2

Question 1: ThIs the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Response to question 1: We have checked our experiment design and original data, meanwhile have added some experiments on volatile components to make our results experiments more complete.

Question 2: Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Response to question 2: Thanks for your careful check. We sought for the help from AJE (American Journal Experts) to refine our manuscript (D3C61K32).

Response to question 2:

Question 3: With the addition of data related to volatile components, the content of the manuscript has now improved to some extent. Related to this, the authors should clarify the method regarding the analysis of volatile components (lines 180-184). Specifically, the modified method, the identification procedure, and the quantification techniques along with key GC-MS parameters should be described. More importantly, it would be great if the authors incorporate the effect of storage time (21 d, 41 d and 61d) on the variations of these volatile molecules (as they did for biogenic amines).

Response to question 3: Thank you for your suggestion. We have added relevant contents such as the determination method, results and discussion of fermented sausage flavor. (line36-37, 48, 185-189, 273-285, 374-375, page 2, 3, 9-10, 15-20, 25).

Attachment

Submitted filename: response to reviewers.docx

Click here for additional data file. (15.8KB, docx)

Decision Letter 5

A M Abd El-Aty

19 Jan 2022

Effect of Collagen Casing on the Quality Characteristics of Fermented Sausage

PONE-D-21-28210R5

Dear Dr. Duan,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

A. M. Abd El-Aty

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #2: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #2: No

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #2: The authors have addressed the issues raised during the past review processes.

The manuscript is now in a good standard and can be considered for publication after rigorous language edition.

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #2: No

Acceptance letter

A M Abd El-Aty

25 Jan 2022

PONE-D-21-28210R5

Effect of collagen casing on the quality characteristics of fermented sausage

Dear Dr. Duan:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

If we can help with anything else, please email us at plosone@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Prof. A. M. Abd El-Aty

Academic Editor

PLOS ONE

Associated Data

    This section collects any data citations, data availability statements, or supplementary materials included in this article.

    Supplementary Materials

    S1 Table. Data of color measurement during processing and storage of fermented sausages.

    (XLSX)

    Click here for additional data file. (9.5KB, xlsx)
    S2 Table. Data of water activity measurement during processing and storage of fermented sausages.

    (XLSX)

    Click here for additional data file. (8.8KB, xlsx)
    S3 Table. Data of pH during processing and storage of fermented sausages.

    (XLSX)

    Click here for additional data file. (8.8KB, xlsx)
    S4 Table. Data of microbiological indicators during processing and storage of fermented sausages.

    (XLSX)

    Click here for additional data file. (9.8KB, xlsx)
    S5 Table. Data of TPA measurement during storage of fermented sausages.

    (XLSX)

    Click here for additional data file. (9.7KB, xlsx)
    S6 Table. Determination results of volatile flavor substances during storage of fermented sausages.

    (XLSX)

    Click here for additional data file. (67.8KB, xlsx)
    S7 Table. Data of biogenic amine measurement during storage of fermented sausages.

    (XLSX)

    Click here for additional data file. (10.7KB, xlsx)
    S8 Table. Basic data of heatmap.

    (XLSX)

    Click here for additional data file. (10.7KB, xlsx)
    Attachment

    Submitted filename: Response to editor.docx

    Click here for additional data file. (49KB, docx)
    Attachment

    Submitted filename: 2nd Response to Reviewers.docx

    Click here for additional data file. (18.1KB, docx)
    Attachment

    Submitted filename: Response to Reviewers.docx

    Click here for additional data file. (15.8KB, docx)
    Attachment

    Submitted filename: response to reviewers.docx

    Click here for additional data file. (15.8KB, docx)

    Data Availability Statement

    All relevant data are within the paper and its Supporting Information files.

    Articles from PLoS ONE are provided here courtesy of PLOS

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