Quality properties of Chinese Sichuan-style sausages as affected by Chinese red wine, yellow rice wine and beer

ABSTRACT

The objective of this research was to investigate quality properties of Chinese Sichuan-style (CSS) sausages treated with Chinese red wine (RW), yellow rice wine (YR) and Chinese beer (CB). The CSS sausages treated with 13% Chinese liquor as the control, and other three groups treated with 6.5% Chinese red wine as the RW group, 6.5% Chinese yellow rice wine as the YR group, and 6.5% Chinese beer as the CB group on the basis of 6.5% Chinese liquor addition were prepared. Results showed that RW, YR and CB group CSS sausages displayed higher L^⁎(39.46–43.44) and A_w, lower pH (5.78–5.91) and protein content (33.75–37.44%) compared with those of the control. The YR group sausages had higher b^⁎ value (23.25), total area% of aldehydes (3.09%) as well as flavor score (6.72), while the CB group CSS sausages showed higher a* value (24.29), total area% of aldehydes (3.94%) and kenones (2.13%), as well as lower free thiol groups (10.88 nmol/mg protein) relative to those of the control group. No significant difference  in overall quality scores were detected between YR and CB group CSS sausages. The PCA showed that RW group CSS sausages achieved similar effects as CB group for volatile, color and sensory properties, which could separate CB/YR group CSS sausages from those of the other two groups. The results of this study indicated that it was applicable to improve quality properties of CSS sausages by using 6.5% Chinese yellow rice wine or beer on the basis of adding 6.5% Chinese liquor during processing.

KEYWORDS:

• Chinese Sichuan-style sausages
• Chinese liquor
• Chinese yellow rice wine
• beer
• quality attributes
• Volatile compounds

Introduction

Chinese Sichuan (CS) sausages, fermented spontaneously for 4–5 weeks at 8–15°C, is one of

the most important Chinese traditional meat products and has become more popular and acceptable toward consumers for distinctive and better flavor and appearance developments.^[1,2] The traditional CS sausages, encased in natural or artificial casings are usually made with raw pork meat, fat, salt, sucrose, red chili powder, Chinese liquor, red rice powder, white pepper and other seasonings.^[3] Nitrites are usually applied to improve sausage flavor and color in CS sausage processing. The spontaneously fermented Chinese Sichuan-style (CSS) sausages, containing higher level of fat and salt, were also susceptible to microorganism contamination and oxidation, which induced product quality deteriorations, safety problems and healthy risks.^[1,3,4]

Numerous efforts have been made to improve quality and safety properties of CS sausage by means of using chitosan-poly‐vinyl alcohol (PVA) antimicrobial packaging and modified casing,^[3,5] adding tea polyphenol, Flos Sophorae and bamboo leaves as antioxidants,^[6,7] as well as inoculating starter culture of various probiotics as flavor, quality, safety enhancer and nitrite replacer.^[1,8] A number of researchers have also investigated the effects of various storage conditions and processing materials on the flavor, safety (microbial community diversity) and other quality changes during CS sausage processing.^[9,10]

The marked association has been reported between wine and sausage production. Sometimes, wine addition could directly influence sausage aroma profile, chemical and sensory properties.^[11] It has been reported that the use of white and red wine as main ingredients, could provide characteristic flavor, inhibit undesirable bacteria growth and replace nitrates/nitrites in sausage production.^[12] Chinese yellow rice wine (Huangjiu), is one of the oldest traditionally brewed wines in China, which produced a large number of nutrients, health active ingredients and complex flavor compounds during brewing process.^[13] Beer, as one of polyphenol-rich foods,^[14] is the most consumed alcoholic beverage worldwide^[15] due to formation of specific sensory, flavor and taste properties.^[16]

While the effect of Chinese red wine, yellow rice wine and beer addition on changes of food quality attributes has been reported. However, negligible similar research reports on CSS sausages are available. This is the first study on the application of Chinese red wine, yellow rice wine and beer addition in production of CSS sausages. We hypothesized the addition of various wine beverage on the basis of Chinese liquor could make positive effects of improving sausage quality properties. The objective of this study was to determine quality changes of CSS sausages treated with other alcoholic beverages like red wine, yellow rice wine and beer on the basis of Chinese liquor addition, thus water activity (A_w), pH, color, texture, lipid and protein oxidation, volatile compounds, sensory attributes were investigated in this research.

Materials and methods

CSS sausage preparation

The CSS sausages were made and fermented spontaneously according to the methods as described by Li et al.,^[10] with slightly modifications. Fresh lean pork M. longissimus dorsi and back fat, were purchased from Chongqing Yonghui supermarket, chopped in a mincing machine (MJ-LZ25Easy225, Midea, China), respectively. The fresh lean pork batter was mixed thoroughly with 20% back fat, 1.6% salt, 0.6% sugar, 1.3% chili powder, 1.7% spice powder, 0.4% red kojic rice, then divided into four batches of about 2 kg each. Chinese liquor, red wine, yellow rice wine and beer were added to the formula randomly: 13% Chinese liquor (ethanol content: 50%, v/v, Jiangjin distillery, Chongqing, China) as the control, 6.5% Chinese red wine (ethanol content: 11%, v/v, COFCO Great wall wine Co., Ltd, Beijing, China) and 6.5% Chinese liquor as the RW group, 6.5% Chinese yellow rice wine (ethanol content: 14%, v/v, Shaoxing Nuerhong wine Co., Ltd, Shaoxing, China) and 6.5% Chinese liquor as the YR group, 6.5% Chinese beer (ethanol content: 3.3%, v/v, Chongqing beer group Co., Ltd, Chongqing, China) and 6.5% Chinese liquor as the CB group for per kg raw meat. After wines and beer addition, the raw CSS sausages were tumbled and stuffed into hog casings with a diameter of 35 mm, soaked in 95°C water for 20 s, and exhausted the air with a needle. During spontaneous fermentation and ripening process, the sausages were hung in a 4°C cold room at 65% r.h. (relative humidity) for 25 days, cooked in an electric steamer at 100°C for 40 min, vacuum-packaged and stored at −20°C until further use.

Chemical analysis

Physicochemical analysis: Levels of moisture, crude fat, crude protein of four group CSS sausages were determined by ISO recommended methods.^[17]

pH: The pH value was determined according to the method of Huang et al. (2022)^[18] with slight modifications. Briefly, CSS sausage samples (2 g) of four groups were grinded, mixed with 20 mL distilled water, and homogenized using a FSH-2 high-speed homogenizer (Changzhou, China) at 10 000 r/min for 1 min, and pH values were read with a PHS-3C pH meter (Shanghai, China).

A_w: A_w values of the CSS sausages were detected at room temperature using a WA-60A A_w meter (LANDTEK, Guangzhou, China).

Color measurements

Color was measured using a Digieye Digital Imaging System (UK, Verivide), calibrated for internal light (D65) before analysis, was used to measure L^⁎ (lightness), a^⁎ (redness/greenness) and b^⁎ (yellowness/brownness) values of 6 points on the surfaces of CSS sausage samples.^[19]

Texture profile analysis

Cylindrical sections of the CSS sausages (diameter = 2 cm; height = 1 cm) were analyzed at room temperature using a texture analyzer (TA.HD Plus Texture Analyzer, Stable Micro Systems, Surrey, England). According to the method of Cruz-Romero et al. (2022).^[20] The CSS sausages were placed in the center of the platform and compressed to 50% of their original height using a 70 kg P50A probe at 5 mm/s, The following parameters were calculated: hardness (kg), resilience (%), cohesiveness, springiness (%), chewiness (kg) and gumminess (kg).

Thiobarbituric acid-reactive substances (TBARS)

The amounts of TBARS in CSS sausages were determined according to the method of Lee et al.^[21] and Dai et al. (2014),^[22] with slightly modifications. Two grams of CSS sausage was blended with 20 mL distilled water, and one milliliter of the meat homogenate was added to 2 mL trichloroacetic acid/thiobarbituric acid (15%TCA/0.375%TBA) and 3 mL 2% butylated hydroxytoluene in a test tube. The sample solution was mixed, heated in a water bath (95°C) for 40 min, and cooled to room temperature for the reaction between malondialdehyde (MDA) and 2-thiobarbituric acid. The absorbance was measured at 538 nm using a Puxi T6-UV-Vis spectrophotometer (Beijing, China), and the amounts of TBARS were expressed as mg of MDA per kg of meat using a molar extinction coefficient of 1.56 × 10⁵ M⁻¹cm⁻¹.

Carbonyl content

The carbonyl content of CSS sausages was detected by reaction with dinitrophenylhydrazone(DNPH) by forming protein hydrazones, according to the method of Chen et al. (2015)^[23] and Dai et al. (2014)^[22] with slight modifications. After extraction of myofibrillar protein (MP) from the sausages, an aliquot (0.5 mL) of MP was reacted with 1 mL of 10 mM DNPH in 2 M HCl at room temperature for 1 h; another 0.5 mL of MP solution was mixed with 1 mL of 2 M HCl (blank). After incubation, the mixture was treated with 1 mL 20% TCA, and centrifuged at 8500 × g for 5 min. The pellets were washed three times with 1 mL of ethyl/ethyl acetate (1:1, v/v), dissolved in 2 mL of 6 M guanidine hydrochloride, the absorbance of the supernatant was read at 370 nm using a Puxi T6-UV-Vis spectrophotometer (Beijing, China). The carbonyl content is expressed as nmol carbonyl/mg protein using an absorption coefficient of 22,000 M⁻¹cm⁻¹ for protein hydrazones after deduction of blank.

Protein-free thiol groups

5,5′-dithiobis (2-nitrobenzoic acid) was used for determination of free thiol groups in CSS sausage proteins according to Dai et al. (2014)^[22] and Martínez Zamora et al. (2021).^[24] The content of free thiol groups was calculated using a molar extinction coefficient of 11,400 M⁻¹cm⁻¹ for 5,5′-dithiobis at this wavelength. Results were expressed as nmol of total thiol groups per milligram of protein.

Volatile compounds

Analysis of CSS sausage volatile compounds was performed by solid-phase micro-extraction-gas chromatography-mass spectrometry (SPME-GC-MS) method described by Belleggia et al. (2022)^[25] with slightly modifications, using Shimadzu GCMS-QP2010 Plus system and a DuraBond DB-WAX capillary column (30.0 m × 250 µm × 0.25 µm, USA). Briefly, CSS sausage meat sample (1 g) was shredded, placed in a 30-mL headspace vial, and the volatile compounds extraction was carried out by injecting a 65-μm PDMS/DVB SPME fiber (Supelco, Bellefonte, PA) into the vial and exposing it to the headspace for 40 min at 45°C. Afterwards, the SPME fiber was desorbed directly into the injection port of the GC at 250°C for 1 min in the spitless mode. The carrier gas was helium with a flow of 0.88 mL/min. The following temperature program of the GC oven was: 40°C (hold 10 min), ramp to 110°C at 5°C/min, then ramp to 240°C at 10°C/min (hold 15 min). The ion source and interface temperature were maintained at 230°C and 250°C, respectively. Electron ionization mass spectra in full-scan mode were recorded at 70 eV electron energy in the range 31–350 amu. Identification of volatile compounds was achieved by comparing mass spectra with the Wiley and NIST libraries (Wiley 7, NIST 05) or standards molecules (for calculating Kovats Index, Supelco 44,585-U, Bellefonte PA, USA). The proportion of each compound was estimated dividing its mean area by the total area of the chromatogram.

Sensory analysis

The sensory evaluation was determined according to the method of Yuan et al.^[26] and Feng et al.^[11] with slightly modifications. Briefly, 18 panelists (consisting of 12 females and 6 males, age 18–40 years) were recruited from the students and staff of Chongqing Chemical Industry Vocational College randomly, served with sliced (approx. 0.5 mm thickness) CSS sausage samples and room temperature water to clean the palate between samples.

The response to each treatment was determined as the mean value of the responses from all the panelists using a Likert scale with scores from 1 (low) to 9 (high). Attributes included color (1 = brown; 9 = light pink), flavor (1 = not detectable; 9 = extremely intense), texture (1 = very mushy; 9 = very firm) and overall quality (1 = bad; 9 = very good).

Statistical analyses

Three replications of CSS sausage samples were used for each analysis, and all data were statistically compared among groups by One-way analysis of variance (ANOVA) and Duncan’s multiple range test with P < 0.05 in Statistical Product and Service Solutions (SPSS) 16.0 system. Principal component analysis (PCA) was performed on SIMCA-P (11.0). Two principal components, PC1 and PC2 were retained to determine treatment scores.

Results and discussion

Physicochemical analysis

The results of moisture, crude fat, protein, pH and A_w analysis are listed in Table 1. After treated with 6.5% Chinese red wine and 6.5% yellow rice wine on the basis of 6.5% Chinese liquor, the moisture content of CSS sausage samples was significant (P < 0.05) higher (30.44%, 31.01%) than that of the control (26.67%). Compared with control CSS sausages, RW, YR and CB group showed significantly (P < 0.05) higher A_w (0.83–0.85), lower pH (5.78–5.91) and crude protein content (33.75–37.44%). Similar results were reported by Feng et al. (2016).^[11] The significant (P < 0.05) decrease in pH values in RW, YR and CB group CSS sausages comparatively to the control was probably due to CO₂ solubilization in these CSS sausages (Pereira et al. 2015).^[27] In accordance with Utrilla et al.,^[28] pH drop was mostly marked in sausages made with Chinese red wine and yellow wine addition, which displayed less moisture loss and higher A_w values.

Table 1. Effect of adding Chinese red wine, yellow rice wine and beer heat on pH and contents of moisture, crude fat, protein, pH and A_w in Chinese Sichuan-style (CSS) sausages.

Treatments	Control	RW group	YR group	CB group
Moisture (%)	26.67 ± 1.10^a	3.44 ± 0.20^b	31.01 ± 0.88^b	28.29 ± 1.91^ab
Crude fat (%)	30.89 ± 2.38	32.28 ± 1.51	33.36±.78	33.29 ± 3.27
Protein (%)	45.82 ± 2.67^b	35.88 ± 1.20^a	33.75 ± 0.69^a	37.44 ± 1.15^a
pH	6.22 ± 0.01^c	5.91 ± 0.02^b	5.88 ± 0.01^b	5.78 ± 0.03^a
Aw	0.79 ± 0.01^a	083 ± 0.01^b	0.85 ± 0.00^b	0.85 ± 0.01^b

Values are means ± SE of three replicates; ^a-c in the same row with different superscripts indicate

significant differences (P < 0.05). Control, the CSS sausages were added by 13% Chinese liquor; RW group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese red wine; YR group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese yellow rice wine; CB group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese beer.

Color analysis

The influences of Chinese red wine, yellow rice wine and beer addition on color attributes in CSS sausages are illustrated in Table 2. Lightness (L^⁎) values were significantly higher in RW, YR and CB group CSS sausages. The CB group CSS sausages showed significantly (P < 0.05) higher redness (a^⁎) values (24.29) relative to control group (22.25), and the yellowness (b^⁎) values (23.25) from YR group CSS sausages were significantly (P < 0.05) higher than those of control group (19.58). Higher a* values obtained in CB group might be likely due to enhanced antioxidation capability of beer, which was used as a “color-stabilizer” in nitrite-free sausage production.^[29,30] This increase in b^⁎ values of YR group may be associated with the addition of Chinese yellow wine itself, which had bright brown color.^[7,31] Higher L* and b* values in sausages may be attributed to slightly higher phenolic contents in Chinese yellow wine.^[7]

Table 2. Effect of adding Chinese red wine, yellow rice wine and beer on instrumental color and texture attributes in CSS sausages.

Treatments	Control	RW group	YR group	CB group
Color attributes
L*	35.56 ± 0.57^a	43.44 ± 0.93^c	41.46 ± 0.38^bc	39.46 ± 0.68^b
a*	22.25 ± 0.13^a	23.07 ± 0.52^ab	22.52 ± 0.80^a	24.29 ± 0.38^b
b*	19.58 ± 0.45^a	21.56 ± 0.79^ab	23.25 ± 1.37^b	21.98 ± 0.51^ab
Texture profiles
Hardness (kg)	76.67 ± 5.93^b	87.60 ± 2.71^b	6.84 ± 3.27^a	74.78 ± 2.23^b
Resilience (%)	17.28 ± 0.91	17.84 ± 0.26	17.06 ± 0.59	16.66 ± 0.59
Cohesiveness	0.61 ± 0.02	0.61 ± 0.01	0.58 ± 0.01	0.59 ± 0.02
Springiness (%)	88.21 ± 0.86	9.48 ± 0.64	89.53 ± 1.26	88.16 ± 2.32
Chewiness (kg)	41.61 ± 3.76^b	48.75 ± 1.96^b	31.65 ± 2.04^a	39.37 ± 3.35^ab
Gumminess (kg)	47.24 ± 4.53^b	53.99 ± 2.26^b	35.37 ± 2.41^a	44.54 ± 2.65^ab

Values are means ± SE of three replicates; ^a-c in the same row with different superscripts indicate

significant differences (P < 0.05). Control, the CSS sausages were added by 13% Chinese liquor; RW group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese red wine; YR group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese yellow rice wine; CB group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese beer.

Texture analysis

Results for texture profile analysis of CSS sausages by addition of Chinese red wine, yellow rice wine and beer are presented in Table 2. In general, values of hardness (60.84 kg), chewiness (31.65 kg) and gumminess (35.37 kg) were significantly lower (P < 0.05) in YR group CSS sausages relative to those of the control and RW group, which was probably due to lower content of meat protein and higher proteolytic activity of MPs in these samples.^[32,33] Unlike the results of treating with 5–10% red wine on the texture profiles of frankfurter sausages,^[11] 6.5% red wine and 6.5% Chinese liquor addition resulted in no difference of hardness, chewiness and gumminess reductions in CSS sausages relative to those treated by 13% Chinese liquor, which may owe to various processing and ripening methods. It was probably expected that more water incorporation into the sausages might improve textural properties by decreasing hardness of sausages treated by 6.5% Chinese yellow wine and 6.5% Chinese liquor.^[34]

TBARS, carbonyl and free thiol groups analysis

TBARS is an indicator of accurately evaluating the degree of lipid oxidation in meat processing and preservation.^[18] No significant difference (P > 0.05) in TBARS values (0.14–0.28 mg MDA/kg meat) was detected in all four group CSS sausages (Figure 1), which agrees with previously reported results.^[35] Protein oxidation is also associated with free thiol loss and carbonyl formation.^[36] Addition of 6.5% Chinese beer to CSS sausages was found to significantly reduce the free thiol content compared to those of the control and RW group, indicating a prooxidative effect of Chinese beer on sausage thiol oxidation. Further, addition of 6.5% Chinese yellow rice wine tended to reduce free thiol groups, but the decrease was not significant (P > 0.05) compared to the control and RW group CSS sausages. Hence, no protective effect of the Chinese beer against free thiol loss in CSS sausages was observed. Similar acceleration of thiol loss was also observed in raw beef patties treated by white grape, apple, green tea extracts, which suggested that some of these protein-phenol reactions had already taken place during processing of sausages.^[37]

Figure 1. Effect of adding Chinese red wine, yellow rice wine and beer on (a) TBARS, (b) carbonyl and (c) protein free thiol groups in Chinese Sichuan-style (CSS) sausages. Values are means ± SE of three replicates measurements. Error bars represent positive standard errors of the mean. Different lowercase letters (a–c) donate significant difference in four treatment group CSS sausages (P < 0.05). Control, the CSS sausages were added by 13% Chinese liquor; RW group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese red wine; YR group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese yellow rice wine; CB group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese beer.

Volatile compounds analysis

Table 3 lists the identified volatiles in all experimental treatments. In briefly, about 75 volatile compounds including 13 hydrocarbons, 20 alcohols, 10 aldehydes, 7 ketones, 5 acids, 13 esters and 5 other aromatics were identified. 18 compounds were common to all of the Chinese sausage products in previous studies.^[38,39]

Table 3. Effect of adding Chinese red wine, yellow rice wine and beer on volatile profiles in CSS sausages.

No	Volatile compounds	KI	CAS	Control	RW group	YR group	CB group
	Terpenes and Alkanes
1	β-Myrcene	958	123-35-3	0.65 ± 0.65	1.26 ± 0.63	0.38 ± 0.34	1.42 ± 0.21
2	α-Terpine	1143	99-86-5	0.22 ± 0.11	0.18 ± 0.14	0.04 ± 0.04	0.07 ± 0.07
3	D-Limonene	1018	7705-14-8	1.15 ± 0.40	1.29 ± 0.48	1.13 ± 0.12	1.47 ± 0.12
4	β-Phellandrene	964	6153-17-9	0.31 ± 0.08	0.33 ± 0.10	0.20 ± 0.10	0.26 ± 0.03
5	trans-.β.-Ocimene	976	3779-61-1	0.30 ± 0.03	0.32 ± 0.04	0.11 ± 0.11	0.25 ± 0.05
6	γ-Terpinene	998	99-85-4	0.59 ± 0.15	0.61 ± 0.11	0.65 ± 0.11	0.57 ± 0.15
7	1,3,6-Octatriene, 3,7-dimethyl-, (Z)-	976	3338-55-4	0.77 ± 0.08	0.87 ± 0.08	0.86 ± 0.05	0.88 ± 0.18
8	(+)-4-Carene	919	5208-49-1	0.07 ± 0.07^a	0.14 ± 0.07^ab	0.16 ± 0.10^ab	0.42 ± 0.15^b
9	Caryophyllene	1494	87-44-5	0.18 ± 0.01	0.16 ± 0.01	0.16 ± 0.01	0.17 ± 0.02
10	Humulene	1579	6753-98-6	0.08 ± 0.04^ab	0.07 ± 0.04^ab	0.00 ± 0.00^a	0.10 ± 0.00^b
11	p-Cymene	1042	9987–6	0.10 ± 0.05^ab	0.19 ± 0.03^b	0.14 ± 0.02^ab	0.04 ± 0.04^a
12	Cyclohexene, 1-methyl-4-(1-methylethylidene)-	1052	138-86-3	0.14 ± 0.07	0.07 ± 0.07	0.14 ± 0.07	0.00 ± 0.00
13	Tetradecane	1413	629-59-4	0.58 ± 0.16^b	0.09 ± 0.09^ab	0.25 ± 0.25^ab	0.00 ± 0.00^a
	Total			5.14 ± 1.51	5.58 ± 1.44	4.20 ± 0.04	5.66 ± 0.72
	Alcohols
14	Ethanol	463	64-17-5	52.05 ± 1.96	44.17 ± 5.72	46.31 ± 3.15	43.27 ± 1.19
15	2,3-Butanediol	743	513-85-9	0.29 ± 0.07^c	0.18 ± 0.04^bc	0.06 ± 0.06^ab	0.00 ± 0.00^a
16	1,6-Octadien-3-ol, 3,7-dimethyl-	1082	126-90-9	8.06 ± 0.59	9.19 ± 1.28	8.62 ± 0.47	9.65 ±0.61
17	1,5,7-Octatrien-3-ol, 3,7-dimethyl-	1072	29957-43-5	0.80 ± 0.06^a	1.14 ± 0.10^b	1.02 ± 0.02^ab	1.23 ± 0.09^b
18	.alpha.-Terpineol	1143	87-55-5	0.48 ± 0.03	0.55 ± 0.08	0.52 ± 0.03	0.56 ± 0.04
19	Geraniol	1228	106-24-1	0.25 ± 0.03	0.29 ± 0.06	0.18 ± 0.03	0.21 ± 0.04
20	p-Cymen-7-ol	1284	53-60-7	0.14 ± 0.01^b	0.11 ± 0.01^ab	0.09 ± 0.01^a	0.13 ± 0.02^ab
21	2-Allylphenol	1203	1745-81-9	0.23 ± 0.01	0.23 ± 0.03	0.21 ± 0.02	0.21 ± 0.01
22	3-Cyclohexen-1-ol, 4-methyl-1-(1-methylethyl)-, (R)-	1137	608522-90-3	0.45 ± 0.04	0.57 ± 0.08	0.49 ± 0.03	0.58 ± 0.05
23	Phytol	2045	150-86-7	0.08 ± 0.01^b	0.06 ± 0.00^a	0.06 ± 0.00^ab	0.07 ± 0.00^ab
24	2 H-Pyran-3-ol, 6-ethenyltetrahydro-2,2,6-trimethyl-	1255	14049-11-7	0.24 ± 0.04	0.28 ± 0.03	0.25 ± 0.03	0.32 ± 0.02
25	2,6-Octadien-1-ol, 3,7-dimethyl-, (Z)-	1228	106-25-2	0.15 ± 0.02	0.18 ± 0.02	0.16 ± 0.02	0.18 ± 0.01
26	2-Cyclohexen-1-ol, 2-methyl-5-(1-methylethenyl)-, cis-	1206	912560-90-8	0.16 ± 0.02	0.18 ± 0.02	0.13 ± 0.01	0.16 ± 0.02
27	Phenylethyl Alcohol	1136	60-12-8	0.07 ± 0.01^a	0.22 ± 0.05^b	0.34 ± 0.01^c	0.19 ± 0.01^b
28	3,7-Octadiene-2,6-diol, 2,6-dimethyl-	1197	51276-34-7	0.09 ± 0.01^a	0.15 ± 0.02^b	0.17 ± 0.02^b	0.20 ± 0.01^b
29	1,6,10-Dodecatrien-3-ol, 3,7,11-trimethyl-, (E)-	1564	1119-38-6	0.19 ± 0.01	0.21 ± 0.03	0.19 ± 0.02	0.22 ± 0.01
30	1,7-Octadiene-3,6-diol, 2,6-dimethyl-	1227	22460-59-9	0.36 ± 0.02^a	0.56 ±  0.06^b	0.56 ± 0.03^b	0.65 ± 0.03^b
31	n-Nonadecanol-1		1454-84-8	0.19 ± 0.03	0.29 ± 0.03	0.27 ± 0.11	0.30 ± 0.10
32	Phenol, 2,6-dimethoxy-4-(2-propenyl)-	1581	6627-88-9	0.11 ± 0.01	0.12 ± 0.01	0.13 ± 0.01	0.12 ± 0.01
33	beta-Eudesmol		473-15-4	0.13 ± 0.00	0.13 ± 0.01	0.13 ± 0.01	0.12 ± 0.01
	Total			64.52 ± 1.56	58.81 ± 4.00	59.89 ± 3.04	58.35 ± 0.39
	Aldehydes
34	Benzaldehyde	982	100-52-7	0.01 ± 0.01^a	0.00 ± 0.00^a	0.00 ± 0.00^a	0.03 ± 0.01^b
35	Benzaldehyde, 4-methoxy-	1171	6515-21-5	0.30 ± 0.03^a	0.57 ± 0.10^b	0.56 ± 0.03^b	0.73 ± 0.08^b
36	Octadecanal	1999	638-66-4	0.03 ± 0.03^a	0.33 ± 0.07^b	0.21 ± 0.08^ab	0.39 ± 0.10^b
37	7-Tetradecenal, (Z)-	1609	65128-96-3	0.10 ± 0.00	0.29 ± 0.04	0.27 ± 0.10	0.32 ± 0.07
38	Citral	1174	5392-40-5	0.04 ± 0.01	0.02 ± 0.00	0.02 ± 0.01	0.02 ± 0.00
39	Hexadecanal	1800	629-80-1	0.42 ± 0.03^a	1.79 ± 0.25^b	1.70 ± 0.28^b	2.04 ± 0.30^b
40	2-Propenal, 3-(2-methoxyphenyl)-	1378	1504-74-1	0.04 ± 0.00^a	0.06 ± 0.00^a	0.05 ± 0.01^a	0.16 ± 0.01^b
41	(1 R)-(-)-Myrtenal	1136	18486-69-6	0.13 ± 0.07	0.02 ± 0.02	0.12 ± 0.03	0.05 ± 0.02
42	Heptadecanal	1800	629-90-3	0.00 ± 0.00^a	0.04 ± 0.00^b	0.04 ± 0.01^b	0.07 ± 0.01^c
43	Tridecanal	1502	10486-19-8	0.00 ± 0.00^a	0.13 ± 0.07^b	0.10 ± 0.02^ab	0.13 ± 0.02^b
	Total			1.07 ± 0.15^a	3.24 ± 0.55^b	3.09 ± 0.50^b	3.94 ± 0.59^b
	Ketones
44	Thujone	1062	471-15-8	0.23 ± 0.02	0.23 ± 0.05	0.34 ± 0.07	0.40 ± 0.14
45	(-)-Carvone	1190	6485-40-1	0.31 ± 0.03	0.33 ± 0.04	0.28 ± 0.02	0.33 ± 0.03
46	2(3 H)-Furanone, 5-ethenyldihydro-5-methyl-	1049	1073-11-6	0.07 ± 0.02^a	0.11 ± 0.02^a	0.08 ± 0.01a	0.18 ± 0.01b
47	2-Propanone, 1-(4-methoxyphenyl)-	1318	122-84-9	0.52 ± 0.01^a	0.74 ± 0.09^b	.76 ± 0.05b	.79 ± 0.05b
48	1-Propanone, 1-(3-methoxyphenyl)-	1318	37951-49-8	0.18 ± 0.00	0.18 ± 0.02	.20±.01	.20±.00
49	4-Hydroxy-2-methylacetophenone	1363	875-59-2	0.09 ± 0.00^a	0.13 ± 0.02^b	.12 ± 0.01ab	.14 ± 0.01b
50	4 H-Pyran-4-one, 2,3-dihydro-3,5-dihydroxy-6-methyl-	1268	28564-83-2	0.00 ± 0.00^a	0.00 ± 0.00^a	.06 ± 0.01b	.09 ± 0.01c
	Total			1.39 ± 0.07^a	1.72 ± 0.23^ab	1.82 ± 0.12ab	2.13 ± 0.14b
	Acids
51	Acetic acid	576	64-19-7	2.85 ± 0.37^b	3.07 ± 0.50^b	2.31 ± 0.36ab	1.38 ± 0.09a
52	Tetradecanoic acid	1769	544-63-8	0.69 ± 0.25^a	1.14 ± 0.07^ab	1.49 ± 0.37ab	1.72 ± 0.19b
53	Pentadecanoic acid	1869	1002-84-2	0.23 ± 0.12	0.32 ± 0.02	.70±.32	.56±.14
54	Oleic Acid	2175	122-80-1	0.41 ± 0.06	0.61±0.07	.90±.41	.75±.25
55	Dodecanoic acid	1580	143-07-7	0.00 ± 0.00^a	0.10 ± 0.05^a	.03 ± 0.03a	.21 ± 0.02b
	Total			4.18 ± 0.72	5.24 ± 0.57	5.23 ± 1.24	4.61±.23
	Esters
56	Benzoic acid, 2-ethylhexyl ester	1259	5444-75-7	0.15 ± 0.02	0.17 ± 0.05	.14±.02	.14±.03
57	Propanoic acid, 2-hydroxy-, ethyl ester	848	7699-00-5	0.15 ± 0.01^a	0.24 ± 0.01^b	.24 ± 0.01b	.35 ± 0.03c
58	Tetradecanoic acid, ethyl ester	1779	124-06-1	0.72 ± 0.07	0.74 ± 0.14	.50±.05	.57±.11
59	Ethyl 9-hexadecenoate	1986	54546-22-4	1.17 ± 0.08	1.10 ± 0.10	.94±.17	.93±.07
60	Ethyl Oleate	2185	111-62-6	1.68 ± 0.24	1.49 ± 0.20	1.14±.23	1.20±.23
61	Geranyl acetate	1352	105-87-3	0.19 ± 0.01	0.20 ± 0.02	.15±.03	.17±.02
62	cis-sesquisabinene hydrate	1523	119238-96-9	0.16 ± 0.01	0.28 ± 0.05	.15±.08	.18±.04
63	Benzoic acid, 4-methoxy-, ethyl ester	1349	94-30-4	0.28 ± 0.01	0.25 ± 0.05	.24±.02	.26±.01
64	Octadecanoic acid, ethyl ester	2177	111-61-5	0.29 ± 0.08	0.28 ± 0.03	.17±.03	.19±.05
65	Hexadecanoic acid, ethyl ester	1978	628-97-7	3.01 ± 0.22	3.40 ± 0.49	2.75±.20	2.98±.27
66	l-(+)-Ascorbic acid 2,6-dihexadecanoate	4765	4218-81-9	1.21 ± 0.74^a	1.64 ± 0.09^ab	2.58 ± 0.94ab	3.83 ± 0.56b
67	Benzeneacetic acid, ethyl ester	1259	101-97-3	0.19 ± 0.02^b	0.00 ± 0.00^a	.00 ± 0.00a	.00 ± 0.00a
68	3,3’−Dimethoxybenzil	2216	40101-17-5	0.34 ± 0.00	0.33 ± 0.03	.31±.02	.33±.03
	Total			9.53 ± 0.64	1.13 ± 1.18	9.32 ± 1.10	11.14±.75
	Other aromatic compounds
69	Acetoin	717	513-86-0	0.97 ± 0.21^ab	1.32 ± 0.19^bc	1.81 ± 0.19c	.57 ± 0.10a
70	Benzenemethanamine, N,N-dimethyl-	1042	103-83-3	0.76 ± 0.09	0.61 ± 0.17	1.27±.60	.74±.03
71	Pyrazine, trimethyl-	1008	14667-55-1	0.18 ± 0.05	0.08 ± 0.01	.09±.07	.11±.03
72	Pyrazine, tetramethyl-	1121	1124-11-4	0.76 ± 0.15^b	0.44 ± 0.05^ab	.62 ± 0.20ab	.26 ± 0.12a
73	Estragole	1172	140-67-0	0.25 ± 0.02^a	0.24 ± 0.01^a	.34 ± 0.02b	.24 ± 0.01a
	Total			2.93 ± 0.19^ab	2.69 ± 0.17^ab	4.13 ± 0.90b	1.92 ± 0.22a

KI: Kovats index relative to C₈-C₂₀ n-alkanes on DuraBond DB-WAX capillary column (30.0 m × 250 µm × 0.25 µm, USA) installed on a gas chromatograph equipped with a mass detector; Values are means± SE of three replicates; ^a-d in the same row with different superscripts indicate significant differences (P < 0.05). Control, the CSS sausages were added by 13% Chinese liquor; RW group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese red wine; YR group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese yellow rice wine; CB group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese beer.

Alcohols have relative high odor thresholds and their contribution to volatile flavor is less than that of other compounds such as aldehydes.^[40] Among the alcohols, ethanol was the major alcohol compound in the CSS sausages, followed by 1,6-octadien-3-ol, 3,7-dimethyl- and 1,5,7-octatrien-3-ol, 3,7-dimethyl- and 1,7-octadiene-3,6-diol, 2,6-dimethyl- and 3-cyclohexen-1-ol, 4-methyl-1-(1-methylethyl)-(R)- and α-terpineol as well as other alcohol compounds, which may be due to the addition of Chinese liquor, red wine, yellow rice wine and beer during CSS sausage spontaneous fermentation and production.^[38] 2,3-butanediol, which was not present in CB group CSS sausages, was detectable in meat sausages of control, RW and YR group. 2,3-butanediol, formed by the reduction of methyl ketones from the α-oxidation of fatty acids, is also detected in fermented jerky.^[41] Although the area% of 1,5,7-octatrien-3-ol, 3,7-dimethyl- and p-cymen-7-ol and phytol and phenylethyl alcohol and 3,7-octadiene-2,6-diol, 2,6-dimethyl- and 1,7-octadiene-3,6-diol, 2,6-dimethyl- was fluctuating in control, RW, YR and CB group CSS sausages, no significant difference (P > 0.05) was detected in total area% of alcohols.

Terpenes and alkanes, derived from thermal oxidation of lipids, could not contribute significantly to meat flavor.^[42] Terpenes, identified in CSS sausages, were major composition of these compounds. Compounds such as β-myrcene, α-phellendren, D-limonene, terpinene, p-cymene were also detected in pepper-treated sausages.^[11,43] No significant difference (P > 0.05) in total area% of hydrocarbons was detected in these four group CSS sausages.

Most of the aldehydes, derived from lipid and protein oxidation, are major contributors to meat flavor due to their lower odor thresholds.^[42] The RW and CB group CSS sausages showed significantly (P < 0.05) higher area% of benzaldehyde, 4-methoxy- and octadecanal and hexadecanal and heptadecanal and tridecanal and total aldehydes relative to those of the control. Benzaldehyde, octadecanal, hexadecanal and heptadecanal were also typical compounds for Istrian dry-cured ham.^[44] The aliphatic and benzaldehyde-based aldehydes may have herbal, floral and almond notes on sausage flavor.^[45]

Small amounts of ketones were detected in all CSS sausages, which are in agreement with the results obtained by Feng et al. (2016).^[11] 4 H-pyran-4-one, 2,3-dihydro-3,5-dihydroxy-6-methyl- could not detected in control and RW group CSS sausages. The YR and CB group CSS sausages exhibited significantly (P < 0.05) higher area% of 2-propanone, 1-(4-methoxyphenyl)- and 4 H-pyran-4-one, 2,3-dihydro-3,5-dihydroxy-6-methyl- than those of the control. In general, the total area% of ketones from CB group CSS sausages was significantly (P < 0.05) higher than those of the control. 2-propanone is responsible for the aroma of blue cheeses and has an intense odor. Acetophenone imparts sweet rose floral odor.^[44,46] The CB group CSS sausages might give better butter flavor due to slightly higher area% of kenones.

Volatile acid compounds make up 4.18–5.24% of the total area of volatiles. Acetic acid, formed by lipid oxidation, amino acid catabolism, lactic acid bacteria and staphylococci fermentation,^[43] was detected in all CSS sausages. The area% of this compound was significantly (P < 0.05) higher in control and RW group CSS sausages compared to those from CB group. Long chain acids such as tetradecanoic acid, pentadecanoic acid, oleic acid and dodecanoic acid do not directly influence meat flavor due to their relative high olfaction thresholds.^[47] No significant difference (P > 0.05) in total area% of acids was detected in these CSS sausages.

Percentages of esters ranged 9.32–11.14% of the total area of volatiles. No significant difference (P > 0.05) in total area% of esters was detected in these CSS sausages. Benzeneacetic acid, ethyl ester was only detected in control group sausages. The CB group CSS sausages had slightly higher (P < 0.05) area% of propanoic acid, 2-hydroxy-, ethyl ester and l-(+)-Ascorbic acid 2,6-dihexadecanoate than those of the control. Most of ethyl esters, formed from ethanol and carboxylic acids by the action of microorganisms, might slightly add fruity aroma notes to sausage flavor.^[44]

Other aromatic compounds, accounted for 1.92–4.13% of the total area of volatiles. The total area% of these compounds in YR group CSS sausages was significantly (P < 0.05) higher than those of CB group. Significant higher areas% of acetoin and estragole were detected in YR group CSS sausages relative to those of the control and CB group. The CB group CSS sausages displayed significantly (P < 0.05) lower area% of pyrazine, tetramethyl- than those of the control. Acetoin and estragole are also the most important aroma substances in star anise and star anise essential oils, which could give special aroma of fennel and sweet taste.^[48]

Sensory evaluation

In the present study, sensory evaluation suggested that no significant difference (P > 0.05) in color and texture scores of all CSS sausages (Figure 2). The addition of 6.5% Chinese yellow rice wine and Chinese liquor significantly (P < 0.05) improved flavor scores and was more preferred by panelists than sausages made with only 13% Chinese liquor, 6.5% red wine and 6.5% Chinese liquor. Also, panelists scored relatively higher (P < 0.05) overall quality for YR (7.5) and CB group (7.61) CSS sausages than those of RW group (6.56). The addition of 6.5% Chinese beer or yellow rice wine might protect sausage sensory characteristics from oxidative degradations. Better sausage quality properties treated with beer or rice wine have also been reported,^[49,50] which was in consistence with our work.

Figure 2. Effect of adding Chinese red wine, yellow rice wine and beer on sensory characteristics (color, flavor, texture and overall quality) in CSS sausages. Values are means ± SE of three replicates measurements. Error bars represent positive standard errors of the mean. Different lowercase letters (a–b) donate significant difference in four treatment group CSS sausages (P < 0.05). Control, the CSS sausages were added by 13% Chinese liquor; RW group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese red wine; YR group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese yellow rice wine; CB group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese beer.

Principal component analysis

Principal component analysis could determine the differences in these four groups of CSS sausages for proximate composition, pH, A_w, color, texture, TBARS, carbonyl and free thiol groups, volatile profile and sensory characteristics, as well as the correlations between these CSS sausages and each variable (Figure 3). The first principal component (PC1) explained 27.4% and the second principal component (PC2) explained 18.3% of the variations (45.7% in total). Control group CSS sausages, which were located in the negative area of PC1 and PC2, were mainly correlated with pH, protein, alcohols and TBARS. RW group CSS sausages were located in the negative area of PC1 and positive area of PC2, which were mainly associated with texture parameters. CB and YR group CSS sausages, which were located in the positive area of PC1, were clustered together and associated with parameters of color, sensory, fat, moisture, A_w, carbonyl and volatile (acids, esters, ketones, aldehydes and other aromatical compounds) formations, confirming that PC1 separated CB/YR group CSS sausages from the other two group sausages on the basis of volatiles, color and sensory profiles. In addition, the sausages treated by CB is necessary for acids and esters formation, whereas those treated by YR is mainly responsible for aldehydes, ketones and other aromatic compounds formation. Therefore, volatile, sensory, color and texture profiles were the main factors to characterize adequately CB/YR and RW group CSS sausages.

Figure 3. Principal component analysis (PCA) for proximate composition, pH, A_w, color, texture, TBARS, carbonyl and protein free thiol groups, volatile and sensory profiles of control, RW, YR and CB group CSS sausages. Control, the CSS sausages were added by 13% Chinese liquor; RW group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese red wine; YR group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese yellow rice wine; CB group, the CSS sausages were added by 6.5% Chinese liquor and 6.5% Chinese beer.

Conclusion

In summary, the addition of Chinese red wine, yellow rice wine and beer could slightly influence quality properties of CSS sausages. These sausages treated with 6.5% Chinese yellow rice wine/Chinese beer and 6.5% Chinese liquor appeared to provide some potential advantages in color, volatile contribution and sensory properties. However, less free thiol groups was obtained in the sausages treated by 6.5% Chinese liquor and beer due to slightly higher protein oxidation. These results suggest that slightly better quality properties could be expected in YR and CB group CSS sausages compared with those of the control, which had lower L* values, area% of volatile aldehydes. These findings indicated that the application of using Chinese yellow wine or beer of Chinese liquor addition might improve quality properties of CSS sausages. Future research might shed light on isolation and identification of core beneficial bacteria strains, bacterial functions and fermentation mechanisms for the quality and safety improvements in CSS sausages processing and preservation by using Chinese yellow rice wine or beer of Chinese liquor.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

This work was mainly supported by Training Project for College Students’ Innovation and Entrepreneurship (HZY202214315004) and the Science and Technology Research Project of Chongqing Municipal Education Commission (KJQN202204057) in Department of Environmental and Quality Inspection, Chongqing Chemical Industry Vocational College, and also supported by the Science and Technology Research Project of Chongqing Municipal Education Commission (KJQN202000817, KJZD-K20220806) in College of Environment and Resources, Chongqing Engineering Research Center for Processing, Storage and Transportation of Characterized Agro-Products, Chongqing Technology and Business University with the technical assistance.