Combined effects of calcium‐alginate coating and Artemisia fragrance essential oil on chicken breast meat quality

Abstract The objective of the present study was to evaluate the effects of calcium‐alginate (CA) containing Artemisia fragrance essential oils (AFEOs) as a potential antioxidant and antimicrobial coating on quality attributes and shelf life of chicken meat throughout keeping period (4°C). Five treatments were produced as follows: T1 (distilled water as control), T2 (2% CA), T3 (2% CA +500 ppm AFEOs), T4 (2% CA +1000 ppm AFEOs), and T5 (2% CA +1500 ppm AFEOs). The chicken meats packaged in polyethylene bags at atmospheric condition and physicochemical, microbiological, and organoleptic properties were assessed at days 1, 4, 8, and 12. There was no remarkable difference in proximate composition (moisture, ash, protein, and fat) of meat samples by treating with CA or AFEOs. The results revealed that CA +AFEOs coating reduced significantly the pH, total volatile base nitrogen (TVB‐N), and thiobarbituric acid reactive substances (TBARS) values and also displayed higher contents of the total phenolic content (TPC) and redness value when compared with control. According to results, 2% CA +1500 ppm AFEOs reduced 58.3 (mg MDA (malondialdehyde)/kg) and 0.63 (mg/100 g) of TBARS and TVB‐N values when compared to control, respectively. The microbiological count showed that CA +AFEOs had a significantly higher inhibitory impact on the total viable count (TVC), coliforms, molds and yeasts. At day 12, 6.89 Log CFU (colony‐forming units)/g was recorded for TVC in 2% CA +1500 ppm AFEOs, which was the lowest overall. This treatment also displayed the reduction of 2.97 Log CFU/g in coliforms and 3.3 Log CFU/g in molds and yeasts in comparison with uncoated samples. The outcomes of pH, TBARS, TPC, color values, microbiological count, and organoleptic properties suggested 2% CA +1500 ppm AFEOs as an efficient coating for quality stability and improving the shelf life of chicken breast meat without negative impact on organoleptic properties.


| INTRODUC TI ON
Chicken meat due to a high amount of protein, moisture, and pH values is a susceptible place for oxidation reactions and microbiological contamination by pathogenic bacteria such as Campylobacter jejuni, Listeria monocytogenes, Salmonella spp., and Escherichia coli which lead to low shelf life (Wickramasinghe et al., 2019;Yaghoubi et al., 2021). However, the main challenge of researchers and also food industries is in improving the quality stability of fresh chicken meat. For extending the shelf life and quality stability of food products, particularly of meat and meat products, edible coatings and films containing plant extracts and essential oils are known as a novel and potential approach (Gutirerrez-García et al., 2021;Javaherzadeh et al., 2020;Karimian et al., 2019;Shahhoseini et al., 2019;Shahosseini et al., 2021). Compared with conventional packaging like modified atmosphere packaging (MAP), vacuum packaging, the edible coatings due to their ability to block carbon dioxide, oxygen, and water vapor from outside and impede the moisture loss, could directly contact the meat to maintain the quality and prolong the shelf life (Xiong et al., 2020).
Polysaccharides like alginate due to particular properties, such as good film-forming, thickening, and gel-producing capability, are utilized widely as biopolymer films or coating components (Draget et al., 2005). Calcium-alginate because of its high biodegradability, availability, and low cost is an efficient alternative for other natural coatings in the meat industry (Hassan et al., 2018). Natural plant extracts and essential oils (EOs) have been widely utilized to improve the shelf life and quality stability of meat and meat products (Bagheri et al., 2016;Javadian et al., 2017;Safari et al., 2018). The genus Artemisia as a perennial plant that belongs to the Asteraceae family (with more than 500 species) and is widely found in Russia, Iran, and their neighboring regions (Bora & Sharma, 2011). Artemisia fragrance with high antibacterial and antioxidant activities (Jaradat et al., 2022;Yaghoubi et al., 2021) is a rich source of 1,8-cineole, α-thujone, β-thujone, and camphor (Jaradat et al., 2022). There are many researches which have reported the potential antioxidant and antimicrobial properties of Artemisia in meat and meat products such as thigh and breast muscles in broilers (Wan et al., 2016), Scomberoides commersonnianus fillets (Farsanipour et al., 2020), and chicken meat . However, the synergistic impacts of calcium-alginate (CA) with Artemisia fragrance essential oils (AFEOs) were not reported. The objective of the present work was to evaluate the preservative impacts of CA coating incorporated with Artemisia fragrance in chicken breast meat by evaluating the microbiological count and quality stability during storage period (4°C).

| Preparation of Artemisia fragrance essential oil (AFEOs)
The AFEOs were prepared by using the Clevenger-type apparatus, 1 L water +400 g dry material of plant subjected to hydrodistillation. The essential oils (EOs) were protected in dark glass bottles, wrapped with aluminum foil around it, and kept in a refrigerator (4°C) until used in chicken samples (1 day).

| AFEOs compounds' isolation
The AFEOs composition was analyzed by using gas chromatographicmass spectrometric (GC-MS) apparatus (Varian, mod. Saturn 2100T, San Fernando, CA, USA). Helium (as the carrier gas; 1 cm 3 /min) and fused-silica capillary column (50 m × 0.22 mm, 0.25μm film thickness) were utilized for the separation of EO components. The temperatures of detector and injector (splitless 20 cm 3 /min) were set at 260 and 280°C, respectively. From 50°C, the oven condition was increased (at the rate of 2°C/min) to 250°C and kept for 1 h. Compared to peak retention time of standard fatty acid methyl ester (FAMEs), the FAMEs of samples were analyzed and the peak area expressed as the compound percentage (Baldino et al., 2017).

| Preparation of chicken meats
Three separate chicken breast meat (skinless, boneless, and weighed between 2 and 2.5 kg) were purchased from a local slaughterhouse (Azar morgh, EA, Iran) during three successive days (with three different preparations) and transferred to laboratory in ice boxes (five treatments ×three times of sampling with three sampling points for each sample). The chicken meat samples were sized 3 × 3 × 3 cm by a sterile knife. Twenty grams of sodium alginate (Keltone LV, ISP, San Diego, CA, USA) and calcium chloride solution (20 µg/ml) were added to an Erlenmeyer flask and reached 1000 ml by using distilled water (60°C), stirred on a heater (60°C) to homogenize the suspension using a magnetic stirrer, and cooled to room temperature before using the meat samples. The meat samples were treated according to: distilled water as control (T1), 2% CA (T2), 2% CA +500 ppm AFEOs (T3), 2% CA +1000 ppm AFEOs (T4), and 2% CA +1500 ppm AFEOs (T5). All meat samples were immersed in produced solutions for 1 h (at 4°C) and finally the samples were drained, packaged in polyethylene bags, and kept at 4°C for the evaluation of proximate composition, pH, color indices, TBARS, TVB-N, total phenolic content (TPC), organoleptic properties, and microbiological count during days 1, 4, 8, and 12 of keeping time.

| Proximate composition and pH value
The ash and moisture content of chicken samples was measured according to AOAC (Association of Official Analytical Chemists) (2006). The Soxhlet extraction technique was utilized for evaluation of the samples' fat content (AOAC, 2006). The protein content of samples was measured by using the Kjeldahl method (AOAC, 2006).
The meat samples' pH was measured by using the Bozkurt and Erkmen (2007) method as follows: the meat samples (10 g) were homogenized in distilled water (100 ml) and assessed with calibrated pH meter (Hanna, Metrohm, Switzerland).

| Determination of the total volatile base nitrogen (TVB-N)
According to Goulas and Kontominas (2005), the meat samples' TVB-N values were analyzed by the Kjeldahl apparatus with a vapor distillation. The results were reported as mg nitrogen/100 g meat samples.

| Total phenolic content (TPC)
The TPC of chicken meat samples was measured according to Liu et al. (2009) by the Folin-Ciocalteu reagent (Merck, Darmstadt, Germany) assay as follows: 100 ml of boiled distilled water and 50 g of meat were mixed together and kept for 20 min at room temperature. The solution was cooled, filtered, and mixed with saturated sodium carbonate solution (5 ml) and Folin-Ciocalteau reagent (2.5 ml). The solution was vortexed and kept (for 1 h) at a dark place. The TPC of samples was measured by ultravioletvisible (UV-vis) spectrophotometer at 700 nm (Hitachi U-3210; Hitachi, Ltd.). The standard curve was prepared by gallic acid (GA) (Merck) and the data were expressed as mg/100 g of gallic acid equivalent (GAE).

| Determination of color values
The lightness (L * ), redness (a * ), and yellowness (b*) of chicken meat surfaces (both internal and external surfaces) were analyzed by a simple digital imaging system in triplicate (Leon et al., 2006). The meats were cut into 3 × 3 × 1 cm thickness for color assessment.
For capturing images, digital camera (16 megapixels) was utilized under suitable light at room temperature (25°C) before instrument calibration with standard plates. Finally, the images were assessed by MATLAB software (The MathWorks, Inc., Natick, MA, USA) for the evaluation of L*, a*, and b* values of meat samples.

| Sensory properties
A total of 72 persons (48 females and 24 males) aged 20-35 years were selected as panelists who had prior experience about chicken breast meat for evaluation of sensory attributes in six booths with 12 panelists for each booth. A randomized (complete) block design was used for estimating the odor, color, freshness, texture, and overall acceptability of meats. The chicken meat samples were individually labeled with stochastic numbers and randomly tested under controlled light and temperature. The odor, color, freshness, and texture were scored according to the hedonic scale (1: really dislike, 5: really like) and the averages of scores reported as the overall acceptability score for each group (Economou et al., 2009;Stone & Sidel, 2004).

| Statistical analysis
The experimental results with five treatments ×four time periods ×three repetition ×three runs were analyzed according to statistical software SAS (v.9; SAS Institute Inc., Cary, NC, USA).
Variance homogeneity and normal distribution had been determined previously (Shapiro-Wilk). The pH, TPC, color indices, TBARS values, TVB-N, organoleptic attributes, and microbiological counts were evaluated through random block design (considering a mixed linear model), including treatments and refrigerated period as fixed effects, and replicate as a random effect. Proximate composition of meat samples including protein, ash, fat, and moisture was also assessed through analysis of variance (ANOVA) (p < .05), followed by Tukey's test. All data in figures and tables were reported as mean values ±SE.

| Gas chromatography-mass spectrometry analysis
The AFEOs volatile chemical compounds are presented in Table 1.

| Proximate composition and pH values
Physicochemical attributes of coated chicken meats with CA containing AFEOs are presented in Table 2. The content of moisture, fat, ash, and protein ranged between 75.69%-75.99%, 1.38%-1.44%, The pH value, which is usually below 6 in fresh meat, can potentially affect the shelf life of meat by changing microbial levels and bacteriostatic function Cullere et al., 2018).
The pH values of samples increased throughout refrigerated time and the rate of this increase was remarkably higher in control

| Determination of thiobarbituricacid reactive substances (TBARS) and total volatile base nitrogen (TVB-N)
One of the most vital indicators for the measurement of lipid oxidation is the TBARS values, which revealed the secondary products (particularly aldehydes) as a result of lipid oxidation (Cai et al., 2014;Sun et al., 2019). The impacts of CA coating in combination with AFEOs are presented in Table 3. Similarly, Alizadeh Behbahani et al. (2017) and Pabast et al. (2018) reported that coating when applied directly to the meat surface may serve as a barrier. Consequently, this reduces the diffusion of oxygen into the meat surface and retards lipid oxidation.
Moreover, high antioxidant properties of AFEOs can also lead to low oxidation throughout keeping period .

| Total phenolic content (TPC)
The natural compounds such as plant extracts and EOs are rich source of phenolic components renowned for their antioxidant,

| Determination of color values
The lightness (L*), redness (a*), and yellowness (b*) of chicken meat samples were remarkably (p < .05) affected by both coating materials and keeping time ( Table 4)

F I G U R E 2
Total phenolic content (TPC) of chicken meat samples treated with calcium-alginate (CA) combined with Artemisia fragrance essential oils (AFEOs) throughout keeping time at 4°C. T1: Control, T2: 2% CA, T3: 2% CA +500 ppm AFEOs, T4: 2% CA +1000 ppm AFEOs, and T5: 2% CA +1500 ppm AFEOs. a-c Different lowercase letters throughout storage indicate significant (p < .05) differences. A-D Different capital letters between meat samples indicate significant (p < .05) differences reaction of phenolic compounds potentially influenced the b* values. However, treated samples with CA +AFEOs and control displayed the lowest and highest amounts of the browning reaction, respectively.

| Microbiological analysis
The results recorded for TVC, coliforms, molds and yeasts count throughout refrigerated period are given in Table 5. At day 1, the Log CFU/g) and control (9.80 Log CFU/g). The coated samples with CA +1500 ppm AFEOs displayed acceptable levels of TVC at day 12. According to Chouliara et al. (2008), the acceptable limitation for TVC in fresh poultry meat is 6 Log CFU/g, while control sam- Because of the special properties of meat such as optimum pH, adequate supply of nitrogenous substances, and high moisture content, this is known as an ideal medium for the proliferation and growth of fungal species. The chicken meats treated with CA +1000 and 1500 ppm AFEOs showed significantly (p < .05) higher inhibitory effects against molds and yeasts throughout storage intervals.
The initial count of molds and yeasts ranged from 1.51 to 1.91 Log CFU/g, which increased significantly (p < .05) and reached 5.72 and 8.75 Log CFU/g for samples treated with CA +1500 ppm AFEOs and control, respectively, at day 12. Molds and yeasts potentially could grow on surfaces of meats and lead to spoilage and negative effects on organoleptic properties and quality stability. The new coatings acted as a barrier and led to a low oxygen concentration on the surfaces of coated samples. Furthermore, the presence of secondary metabolites in EOs can retard or inhibit the growth of yeasts and molds and bacteria.

| Sensory properties
Sensory attributes, which highly affected the marketing of products, reflect consumers' preference and the overall quality of products to a certain extent. With regard to Figure 3,   , treated lamb with chitosan coating with Satureja plant essential oil (Pabast et al., 2018), and chitosan-gelatin coating +tarragon EOs in pork slices (Zhang et al., 2020).
The outcomes of the present work indicated that the CA coating combined with AFEOs had no adverse effects on the sensory attributes of chicken breast meat. Furthermore, based on the sensory results from microbial growth and chemical reactions, the application of AFEOs in the present work on different aspects of chicken breast meat against microbial spoilage, lipid oxidation, F I G U R E 3 Sensory properties of chicken meat coated with calcium-alginate +AFEOs (Artemisia fragrance essential oils) throughout keeping time at 4°C. T1: Control, T2: 2% CA, T3: 2% CA +500 ppm AFEOs, T4: 2% CA +1000 ppm AFEOs, and T5: 2% CA +1500 ppm AFEOs. a-d Different lowercase letters throughout storage indicate significant (p < .05) differences A-B Different capital letters between meat samples indicate significant (p < .05) differences off-odor, texture, and discoloration throughout the keeping time was acceptable.

| CON CLUS ION
The results for chemical composition and microbiological count revealed that CA coating containing AFEOs on chicken breast meat can lead to good quality properties, enhancement of microbiological safety, and improvement of shelf life throughout keeping time. All treatments declined remarkably microbial counts when compared to control. The quality attributes (TBARS, TVB-N) of treated samples remained within the acceptable range for a longer period. Alginate coating containing 1500 ppm AFEOs had the highest inhibitory effect on lipid and protein oxidations and against microbial growth during keeping time. The outcomes of the present work indicated that the shelf life of chicken breast meat could be remarkably increased by calcium-alginate coating +1500 ppm AFEOs which can be suggested as potential coating materials. According to results obtained from the commercialization of CA and other coating materials such as chitosan, due to the disproportionate price with the application, these compounds can be used in combination with gelatin protein to coating chicken meat in the meat industry and retail.

ACK N OWLED G EM ENT
Thanks to Islamic Azad University of Tabriz for supporting this manuscript.

CO N FLI C T O F I NTE R E S T
The authors reported no potential conflict of interest.

E TH I C A L A PPROVA L
This article does not cover any human or animal studies conducted by any of the authors.

DATA AVA I L A B I L I T Y S TAT E M E N T
Data are available upon request from the authors.