Abstract
The objective of this study was the incorporation of a water–oil (W/O) nanoemulsion for the partial substitution of pig fats and the addition of antioxidant compounds in an emulsified meat system (EMS). The nanoemulsion was formulated with orange essential oil and cactus acid fruit (xoconostle). The treatments were different percentages (0, 1, 2, 3, 4, and 5%) of the nanoemulsion for the substitution of pig fat in the EMS. The proximal analysis (moisture, protein, fat, and ash), texture profile (hardness, cohesiveness, springiness, and chewiness), phenolic compounds and antioxidant capacity 2, 2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-Azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS), and 2-thiobarbituric acid reactive substances (TBARS) were evaluated. All variables showed significant differences (p < 0.05). The results for protein, fat, and ash exhibited increments with the addition of the nanoemulsion, and moisture loss was reduced. The profile showed increments in hardness and chewiness. The addition of the nanoemulsion incremented the phenolic compounds and antioxidant capacity (DPPH and ABTS), decreased production of Malonaldehyde, and reduced lipid oxidation. The result of the addition of the nanoemulsion in the EMS is a product with a substantial nutritional contribution, antioxidant capacity, and excellent shelf life.
Keywords: xoconostle, phenols, ABTS, DPPH, TBARS
1. Introduction
In the meat industry, fat is very important for emulsified products. Fat is responsible for emulsion stability and water retention capacity, further providing energy, essential fatty acids, and fat soluble vitamins [1]. The fat soon degrades due to oxidation, thereby producing poor sensory characteristics, discoloration, and rancidity [2]; in addition, fat oxidation results in a reduction in shelf life and production of toxic compounds [3].
Emulsified meat products are enhanced with synthetic antioxidant compounds, including butyl-hydroxytoluene, butyl-hydroxyanisole, and t-butyl-hydroxyquinone, among others, to reduce fat oxidation and extend shelf life [4]. These synthetic products have the disadvantages of promoting toxicological, mutagenic, and carcinogenic effects [5,6,7]. One alternative option is the use of natural antioxidants in meat products [2].
The cactus acid fruit, xoconostle, from the genus Opuntia, contains phenolic compounds, carotenoids, betacyanins, and betalains [8]. These bioactive compounds have shown antioxidant activity [9] and antibacterial activity [10]. Furthermore, the orange essential oils include terpenes, such as D-limonene, that protect fat against oxidizing compounds [11]. In addition, essential oils have been shown to possess antibacterial, antifungal, and antioxidant activities [12,13], so these components can be used as functional ingredients in foods [14,15]. The antioxidant compounds are sensitive to external factors, such as light, temperature, and oxygen. One way to protect these compounds is by using encapsulation, such as in the form of nanoemulsions. This type of encapsulation has the advantage of improving the transportation and controlling the release of active molecules through the biological membrane [16].
Sharma et al. [17] incorporated four types of essential oils (clove, holy basil, cassia, and thyme) in emulsified chicken and demonstrated a reduction in fat oxidation. Wang et al. [3] substituted pig fat with camellia oil gel in sausage and found favorable results, such as reduced fat, lower moisture, and minor values of 2-thiobarbituric acid reactive substances (TBARS).
The objective of this work was the partial substitution of pig fat with water-oil W/O emulsions containing cactus acid fruit (xoconostle) in an emulsified meat system to evaluate the physicochemical characteristics, texture profile, and oxidative stability for 60 days. The objective of this work was to evaluate the effect of the partial substitution of pig fat with W/O emulsions containing cactus acid fruit (xoconostle) in an emulsified meat system on physicochemical characteristics, texture profile, and oxidative stability.
2. Materials and Methods
2.1. Preparation of the Nanoemulsion
The nanoemulsion was water in oil (W/O). It was prepared according to the methodology of Guler et al. [18] with some modifications. The continuous phase was orange essential oil (Hilmar Ingredients, USA) (70%), the dispersed phase was the cactus acid fruit (xoconostle) (10%), and the surfactant was liquid soya lecithin (Hilmar Ingredients, USA) (20%). All components were stirred using an ultrasonic processor (Ultrasonic Sonics Vibra-Cell, VCX 130, USA). A 6 mm probe was used and 20 intervals (20 s/interval) of sonication and recesses of 10 s were established to obtain the necessary drop size. The ultrasonic processor was used at 80% amplitude at a frequency of 20 kHz, and the mixture was placed in an ice bath to avoid temperature increases during mixing. The droplet size distribution of the nanoemulsion was determined with the dynamic laser light scattering technique using Zetasizer equipment (Nano-ZS2000 Model Malvern Instruments Ltd. Malvern, Worcestershire, United Kingdom) by placing the sample in a glass cell. Five replicates were considered for each formulation [19]. Furthermore, the phenolic content and antioxidant activity (2, 2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-Azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS)) were determined in the nanoemulsions.
2.2. Production of the Emulsified Meat System
The emulsified meat system was carried out according to the method described by Cofrades et al. [20] with some modifications. Formulations with different percentages of animal fat and nanoemulsion were made (Table 1). The minced meat (1 cm2), salt, and ice were placed in a cutter (Dito-Sama F 23200 GBR, Aubusson, France) and beaten for two minutes, and then nanoemulsion was added to the mixture and beaten for one minute more.
Table 1.
Formulation of emulsified meat systems with different percentages of nanoemulsion.
| Treatments | Meat % | Fat % | Nanoemulsion % | Ice % | Salt % |
|---|---|---|---|---|---|
| 0% | 65 | 20 | 0 | 13 | 2 |
| EMSN 0% | 65 | 19 | 1 | 13 | 2 |
| EMSN 2% | 65 | 18 | 2 | 13 | 2 |
| EMSN 3% | 65 | 17 | 3 | 13 | 2 |
| EMSN 4% | 65 | 16 | 4 | 13 | 2 |
| EMSN 5% | 65 | 15 | 5 | 13 | 2 |
Emulsified meat system with nanoemulsion (EMSN).
The fat was incorporated into the mixture, maintaining a temperature no higher than 16 °C. The mixture was fed through a stuffing device (BG-PRUFRZERT Inc., City of México, México) and injected into 20 mm diameter synthetic cellulose casings (Viscofan Brand Inc., City of México, México). The filled casings were heated to 72 °C for 30 min, and then subjected to thermal shock by being placed in ice. Finally, the meat emulsion in casings were vacuum packed in bags (Zubex Inc., City of México, México) in a sealer (Tor Rey EVD48, City of México, México) and refrigerated at 4 °C.
2.3. Proximal Composition
The proximal analysis was performed according to the official methods of the Association of Official Agricultural Chemists (AOAC) edited by Horwitz [21]. The moisture was calculated by drying a sample in a stove at 100 °C for 8 h (Official Method 925.09), the fat content by the Soxhlet method (Official Method 923.05), the ash percentage was determined by the incineration of the muffle samples at 550° C for 8 h (Official Method 923.03), and the protein content by the Kjeldahl method (Official Method 981.10).
2.4. Texture Profile Analysis (TPA)
These tests were performed according by Cofrades et al. [22] with some modifications. Eight repetitions were performed for each treatment. Cubes of 1 × 1 × 1 centimeters were elaborated and a texturometer (Brookfield CT3 texture analyzer, Brookfield Engineering Laboratories, Inc. Middleboro, MA, USA) was used. The samples were axially compressed to 50% of their original height with a 4.5 kg load cell at a speed of 1 mm/s, with the use of a TA3/1000 probe and a TA-BT-KI table. The parameters measured were hardness, cohesiveness, springiness, and chewiness. The test was performed at room temperature.
2.5. Total Phenols
The content of total phenols was done following a modified version of the methodology by Singleton et al. [23]. The samples were diluted to 1:10. Then, 0.5 mL of sample was mixed with 2.5 mL of previously diluted (1:10) Folin-Ciocalteau reagent (Sigma-Aldrich, St. Louis, MO, USA) and 2 mL of 7.5% sodium carbonate (Fermont) was added. The mixture was left for 120 min in total darkness. After, the samples were read in a spectrophotometer (JENWAY 6715 Ultraviolet/Visible (UV/V), Staffordshire, UK) at a wavelength of 760 nm. The results were expressed as mg of gallic acid equivalents for 100 g of emulsified meat system with nanoemulsion (EMSN) (GAE/100 g of EMSN).
2.6. DPPH
The methodology of Brand-Williams et al. [24] for DPPH test was used with some modifications. Here, 0.0039 g of DPPH (2,2-diphenyl-1-picrylhydrazyl) (Sigma-Aldrich, USA) in 50 mL of 80% methanol (JT Baker, VWR International. Tultitlán, México) was mixed and left for 2 h in the dark, then calibrated at 0.7 ± 0.1 absorbance. Then, 0.5 mL of this mixture was added to 2.5 mL of DPPH solution and left in darkness for 60 min. After, the samples were read at 517 nm in a spectrophotometer (JENWAY 6715 UV/V, UK). The results were expressed as mg of ascorbic acid equivalents for 100 g of emulsified meat system with nanoemulsion (EMSN) (AAE/100 g of EMSN).
2.7. ABTS
Here, 7 mM (10 mL) of 2, 2′-Azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) (Sigma-Aldrich, St. Louis, MO, USA) and 2.45 mM (10 mL) of potassium persulfate were mixed. The mixture was left in complete darkness for 16 h. Next, the mixture was adjusted with 20% ethanol to obtain a value of 0.7 ± 0.1 absorbance. The final solution (3.9 mL) was taken and 100 μL of sample was added. The mixture was read at 734 nm [25]. The results were expressed as mg of ascorbic acid equivalents for 100 g of emulsified meat system with nanoemulsion (EMSN) (AAE/100 g of EMSN).
2.8. 2-thiobarbituric acid reactive substances (TBAR)
Lipid oxidation was evaluated according to Wang et al. [26] with some modifications. An extractor solution was prepared containing 7.5 % trichloroacetic acid (Fermont PA Cert, Monterrey, México), 0.1% gallic acid (Fermont PA Cert, Monterrey, México), and 0.1 % EDTA, disodium salt dehydrate (Baker ACS, México). Then, 2.5 g of samples were taken and homogenized with 25 mL of extractor solution in an Ultraturrax T25 (IKA-Werke GmbH & Co. KG) 3000 rpm for 1 min. The homogenate was centrifuged at 6000× g forces at 20 °C for 10 min. The supernatant (2 mL) was mixed with 80 mM (2 mL) of thiobarbituric acid (BP 50067 lllkirch, Strasbourg, France) (TBA). The mixture was incubated at 40 ° C for 90 min and it was read at 532 nm. The TBARS values were interpreted with the calibration curve of 1,1,3,3-tetramethoxypropane (Malonaldehyde) (Sigma-Aldrich, St. Louis, MO, USA) in different concentrations and the results were expressed in milligrams of malonaldehyde (MDA)per kilogram of sample (mg MDA /Kg).
2.9. Statistical Analysis
The experimental design was completely random. The results were analyzed by ANOVA, when there were significant differences (p < 0.05), comparison of media (Tukey) was used with the statistical program STATGRAPHICS C. XVI Version 16.1.03 (Statgraphics Technologies Inc., The Plains, VA, USA).
3. Results and Discussion
3.1. Nanoemulsions and Characterization
The drop diameter was 73 ± 6 nm and the Z potential value was −107 mV. Both parameters are characteristic of nanoemulsions [27,28]. Our results are similar to those reported in Gago et al. [29] for nanoemulsions of clove and lemongrass essential oil. The phenolic content was 184.3 mg GAE/100 g, the antioxidant activity from DPPH was 97.76 mg AAE/100 g, and ABTS was 126.3 mg AAE/100g in the nanoemulsions.
3.2. Proximate Composition
Significant differences (p < 0.05) were observed in the moisture of the meat emulsion system in the different treatments and times. Treatments with the nanoemulsions demonstrated a reduced loss of moisture (Table 2), and similar results were reported by Sharma et al. [17] in chicken sausage with the addition of different essential oils. The major reason for the retention of water could be that the nanoemulsions contain soy lecithin in the formulation, which was used as an emulsifier [30,31].
Table 2.
Proximal composition of the emulsified meat system with nanoemulsion for the parameters of moisture, protein, fat and ash.
| Days | EMSN 0% | EMSN 1% | EMSN 2% | EMSN 3% | EMSN 4% | EMSN 5% | |
|---|---|---|---|---|---|---|---|
| Moisture | 1 | 68.58 ± 0.085 aC | 68.53 ± 0.007 aB | 68.52 ± 0.019 aD | 68.59 ± 0.094 aB | 68.65 ± 0.093 aE | 68.67 ± 0.092 aD |
| 15 | 67.88 ± 0.018 aC | 67.94 ± 0.479 aB | 67.98 ± 0.099 aC | 67.92 ± 0.382 aB | 67.98 ± 0.013 aD | 67.96 ± 0.112 aC | |
| 30 | 65.14 ± 0.427 aB | 65.60 ± 0.107 aA | 65.72 ± 0.077 aB | 65.89 ± 0.036 aA | 66.05 ± 0.022 aC | 66.11 ± 0.083 aB | |
| 45 | 64.46±0.252 aAB | 65.10 ± 0.075 bA | 65.23 ± 0.001 bA | 65.32 ± 0.001 bcA | 65.67± 0.009 cdB | 65.80 ± 0.003 dA | |
| 60 | 63.58 ± 0.002 aA | 64.95 ± 0.002 bA | 65.13 ± 0.010 cA | 65.24 ± 0.011 dA | 65.40 ± 0.017 eA | 65.58 ± 0.009 fA | |
| Protein | 1 | 14.89 ± 0.020 aA | 15.09 ± 0.008 abA | 15.10± 0.024 abA | 15.15±0.009 abcA | 15.29± 0.146 bcA | 15.40 ± 0.103 cA |
| 15 | 15.39±0.041 aB | 15.47±0.199 aAB | 15.53 ± 0.073 aB | 15.63 ± 0.024 aB | 15.67 ± 0.306 aA | 15.83 ± 0.056 aB | |
| 30 | 15.49± 0.022 aBC | 15.54 ± 0.031 aB | 15.61 ± 0.089 aB | 15.84 ± 0.017 bC | 15.87± 0.037 bAB | 16.00 ± 0.035 bB | |
| 45 | 15.55 ± 0.005 aC | 16.08 ± 0.066 bC | 16.24 ± 0.023 cC | 16.36 ± 0.024 cdD | 16.41± 0.054 dBC | 16.46 ± 0.035 dC | |
| 60 | 16.15 ± 0.059 aD | 16.65 ± 0.003 bD | 16.69 ± 0.002 bD | 16.84 ± 0.008 cE | 16.89 ± 0.008 cC | 16.94 ± 0.012 cD | |
| Fat | 1 | 8.83 ± 0.058 aA | 9.39 ± 0.007 bA | 10.26 ± 0.056 cA | 10.60 ± 0.077 dA | 11.74 ± 0.012 eA | 12.23 ± 0.092 fA |
| 15 | 9.35 ± 0.186 aB | 10.11 ± 0.037 bA | 10.87 ± 0.051 cA | 11.12 ± 0.070 cB | 12.44 ± 0.147 aB | 12.61 ± 0.192 dA | |
| 30 | 13.38 ± 0.024 aC | 14.29 ± 0.257 abB | 14.43 ± 0.370 bB | 14.75 ± 0.026 bC | 14.90 ± 0.322 bC | 15.32 ± 0.325 bB | |
| 45 | 13.95 ± 0.061 aD | 14.53 ± 0.646 abB | 14.59 ±0.087 abB | 14.81± 0.008 abC | 15.21 ± 0.099 bC | 15.29 ± 0.015 bB | |
| 60 | 14.52 ± 0.008 aE | 14.64 ± 0.068 abB | 14.78 ± 0.022 bB | 14.95 ± 0.011 dA | 15.33 ± 0.012 dC | 15.69 ± 0.024 eB | |
| Ash | 1 | 1.94 ± 0.037 aA | 1.94 ± 0.017 aA | 1.94 ± 0.001 aA | 1.94 ± 0.005 aA | 1.95 ± 0.001 aA | 1.95 ± 0.006 aA |
| 15 | 1.95 ± 0.002 aA | 1.96 ± 0.001 abAB | 1.96 ± 0.002 abcA | 1.97 ± 0.003 bcB | 1.97 ± 0.002 cB | 1.97 ± 0.002 cB | |
| 30 | 1.96 ± 0.001 aA | 1.96 ± 0.002 aAB | 1.96 ± 0.007 aB | 1.97 ± 0.001 abB | 1.98 ± 0.001 abB | 1.98 ± 0.003 bB | |
| 45 | 1.97 ± 0.007 aA | 1.97 ± 0.007 aAB | 1.97 ± 0.002 aBC | 1.97 ± 0.001 aB | 1.98 ± 0.001 aB | 1.98 ± 0.003 aB | |
| 60 | 1.98 ± 0.002 aA | 1.98 ± 0.006 aB | 1.98 ± 0.001 aC | 1.98 ± 0.004 aB | 1.98 ± 0.006 aB | 1.98 ± 0.003 aB |
Emulsified meat system with nanoemulsion (EMSN). The lowercase letters in the superscript indicate significant differences (p < 0.05) between treatments (rows), and uppercase letters indicate significant differences in each treatment with respect to time (columns) (p < 0.05).
Significant differences (p < 0.05) were observed in protein between the different treatments and times. The major protein content was observed in the treatments with nanoemulsions (Table 2). Choi et al. [1] found similar results in the substitution of pig fat with vegetable oil in the emulsified meat system. However, Bolger, Brunton, and Monahan [32] did not find significant differences (p > 0.05) in protein content in an emulsified product with encapsulated flaxseed oil. The increment in protein could be due to soy lecithin, which contains amino acids.
The EMSN showed a significant increment (p < 0.05) in the content of fat after the addition of the nanoemulsion (Table 2). In contrast, Choi et al. [1] found less fat with the addition of vegetable oils in EMS; however, the quality of the lipid provided by the nanoemulsion is better compared to that provided by pig fat. The orange essential oil contains antioxidant compounds, such as D-limonene, according to Chasquibol et al. [11].
The values of ash were between 1.94 and 1.95 in the treatment with EMSN 5% (Table 2). Choi et al. [1] reported similar results (1.72 to 1.97) with the addition of vegetables oils in EMS.
3.3. Texture Profile Analysis (TPA)
The nanoemulsion significantly (p < 0.05) affected the hardness of the EMSN. Treatment with the 5% nanoemulsion produced the most substantial hardness (Table 3). Similar results were reported by Youssef and Barbut [33] in a meat batter with canola oil. These authors attributed the increase in the hardness to the oil’s smaller globule size and the enhanced interaction between proteins.
Table 3.
Texture profile analysis (TPA) for the parameters hardness, cohesiveness, springiness, and chewiness in the emulsified meat system with nanoemulsion.
| Days | EMSN 0% | EMSN 1% | EMSN 2% | EMSN 3% | EMSN 4% | EMSN 5% | |
|---|---|---|---|---|---|---|---|
| Hardness (N) | 1 | 12.49 ± 0.344 bA | 12.38 ± 0.307 bA | 12.63 ± 0.302 aA | 13.44± 0.358 cA | 14.11 ± 0.306 dA | 14.57± 0.333 dA |
| 15 | 13.68 ± 0.238 bB | 12.93 ± 0.357 aB | 12.94 ± 0.406 aB | 14.52 ± 0.270 cB | 15.16 ± 0.254 dB | 15.53 ± 0.252 dB | |
| 30 | 14.40 ± 0.238 bC | 13.51 ± 0.336 aC | 14.13 ± 0.374 aB | 15.43 ± 0.245 cC | 16.57 ± 0.332 dC | 16.47 ± 0.406 dC | |
| 45 | 15.12 ± 0.681 cD | 13.73 ± 0.165 aC | 14.94 ± 0.373 bC | 16.43 ± 0.261 dD | 17.58 ± 0.313 eD | 17.50 ± 0.288 eD | |
| 60 | 17.76 ± 0.252 cE | 14.59 ± 0.318 aD | 15.25 ± 0.356 bD | 17.58 ± 0.314 cE | 18.40 ± 0.264 dE | 18.52 ± 0.299 dE | |
| Cohesiveness | 1 | 0.65 ± 0.007 abC | 0.65 ± 0.005 abC | 0.64 ± 0.005 aD | 0.65 ± 0.006 abC | 0.64 ± 0.004 abD | 0.65 ± 0.005 bC |
| 15 | 0.64 ± 0.004 bC | 0.63 ± 0.011 abB | 0.63 ± 0.007 aC | 0.63 ± 0.007 abB | 0.63 ± 0.007 abC | 0.63 ± 0.006 abB | |
| 30 | 0.63 ± 0.005 abB | 0.63 ± 0.005 bB | 0.62 ± 0.009 aBC | 0.63 ± 0.004 abB | 0.63 ± 0.006 abBC | 0.63 ± 0.007 bB | |
| 45 | 0.62 ± 0.007 abB | 0.61 ± 0.014 abA | 0.61 ± 0.010 aAB | 0.62 ± 0.007 abA | 0.62 ± 0.004 bB | 0.62 ± 0.005 abA | |
| 60 | 0.60 ± 0.011 aA | 0.61 ± 0.010 abA | 0.61 ± 0.009 abA | 0.61 ± 0.007 abA | 0.61 ± 0.008 abA | 0.62 ± 0.006 bA | |
| Springiness (mm) | 1 | 4.36 ± 0.029 bA | 4.33 ± 0.020 abC | 4.34 ± 0.021 abC | 4.32 ± 0.031 abD | 4.31 ± 0.024 aB | 4.31 ± 0.039 aB |
| 15 | 4.35 ± 0.020 dB | 4.26 ± 0.014 aB | 4.33 ± 0.021 cdC | 4.30 ± 0.024 bcCD | 4.29 ± 0.024 bAB | 4.29 ± 0.015 bAB | |
| 30 | 4.34 ± 0.018 cB | 4.24 ± 0.017 aB | 4.32 ± 0.019 cBC | 4.27 ± 0.027 abBC | 4.28 ± 0.023 abAB | 4.28 ± 0.025 bAB | |
| 45 | 4.26 ± 0.023 abCA | 4.23 ± 0.033 aB | 4.30 ± 0.019 cAB | 4.24 ± 0.036 abAB | 4.26 ± 0.029 abcA | 4.27 ± 0.031 bcAB | |
| 60 | 4.24 ± 0.024 bcA | 4.18 ± 0.031 aA | 4.28 ± 0.028 cA | 4.21 ± 0.053 abA | 4.25 ± 0.040 bcA | 4.26 ± 0.030 cA | |
| Chewiness (NXmm) | 1 | 33.41 ± 1.25 aA | 34.15 ± 0.887 bA | 36.95 ± 0.543 bA | 37.95± 0.358 cA | 38.43 ± 0.990 cA | 38.55± 0.525 cA |
| 15 | 34.64 ± 1.04 aA | 35.24 ± 0.792 aB | 37.96 ± 0.921 bA | 38.97 ±0.542 bcA | 39.29 ±0.831 cAB | 41.01 ± 0.752 dB | |
| 30 | 37.29 ± 1.04 bB | 35.88 ±0.984 abC | 42.00 ± 0.664 dB | 41.64 ± 0.877 dB | 40.14 ± 0.981 cB | 42.72 ± 0.771 dC | |
| 45 | 38.78 ± 0.84 bC | 36.23 ± 0.900 aC | 43.33± 0.928 cC | 42.63 ± 0.850 cB | 42.05 ± 0.870 cC | 43.29 ± 0.880 cC | |
| 60 | 41.26 ± 0.775 bD | 37.23 ± 0.839 aD | 45.98 ± 0.988 cD | 45.31 ± 0.652 cC | 45.15 ± 0.837 cD | 46.25 ± 0.904 cD |
Emulsified meat system with nanoemulsion (EMSN). The lowercase letters in the superscript indicate significant differences (p < 0.05) between treatments (rows). and uppercase letters indicate significant differences in each treatment with respect to time (columns) (p < 0.05).
The nanoemulsion did not affect the cohesiveness of the EMSN (Table 3). Wang et al. [3] reported the same results after the partial substitution of pig fat with camellia oil gel in sausage. In contrast, Choi et al. [1] observed an increment after the addition of vegetable oils in an EMSN.
No significant differences (p > 0.05) were observed between treatments with respect to springiness (Table 3). The incorporation of flaxseed oil did not affect the springiness of chicken sausage [32]. The EMSN did not show changes in springiness due to the addition of oils or nanoemulsions.
The EMSN exhibited a significant increment (p < 0.05) in chewiness after the incorporation of the nanoemulsion (Table 3). These results coincide with those reported by Youssef and Barbut [33], Choi et al. [1], and Bolger et al. [32] with respect to the substitution of fat with vegetable and seed oils in EMSN. The increase in chewiness is related to the protein incorporated within the nanoemulsion.
The effect on shelf life exhibited significant differences (p < 0.05) after the substitution of pig fat with the nanoemulsions. The EMSN showed increased hardness and chewiness but reduced cohesiveness and springiness, and these effects could be attributed to the loss of moisture during storage.
3.4. Total Phenols and Antioxidant Activity
The contents of phenols were significantly enhanced (p < 0.05) by the incorporation of the nanoemulsions (Table 4) because the nanoemulsions contain phenolic compounds from the xoconostle extract. The addition of cherry extract to sausage also increased the content of phenols [34].
Table 4.
Phenols, antioxidant activity via the DPPH and ABTS methods, and oxidative stability via the TBARS method in an emulsified meat system with nanoemulsion.
| Days | EMSN 0% | EMSN 1% | EMSN 2% | EMSN 3% | EMSN 4% | EMSN 5% | |
|---|---|---|---|---|---|---|---|
| Phenols mg GAE/100g | 1 | ND | 12.76 ± 0.345 aC | 13.29 ± 0.486 aC | 15.64 ± 0.177 bD | 19.86 ± 0.215 cD | 24.93 ± 0.170 dE |
| 15 | ND | 12.22 ± 0.385 aC | 14.39 ± 0.049 bD | 14.47 ± 0.098 bC | 15.21 ± 0.098 cC | 18.09 ± 0.161 dD | |
| 30 | ND | 11.25 ± 0.098 aB | 12.31 ± 0.078 bB | 12.39 ± 0.345 bB | 15.10± 0.085 cC | 16.04 ± 0.085 dC | |
| 45 | ND | 10.45 ± 0.274 aA | 11.71± 0.148 aAB | 12.50 ± 0.098 bB | 13.25 ± 0.090 cB | 14.58 ± 0.085 dB | |
| 60 | ND | 10.40 ± 0.098 aA | 11.39 ± 0.098 bA | 11.56 ± 0.177 bA | 11.76 ± 0.085 bcA | 12.16 ± 0.177 cA | |
| DPPH mg AAE/100g | 1 | 15.55 ±0.288 aC | 18.13 ± 0.377 bD | 19.20 ± 0.108 cD | 19.26 ± 0.188 cD | 19.76 ± 0.288 cD | 19.89 ± 0.288 cD |
| 15 | 13.60 ± 0.188 aC | 17.88 ± 0.474 bD | 19.01 ± 0.474 cD | 18.69 ± 0.188 cD | 19.07±0.499 cdCD | 19.89 ± 0.288 dD | |
| 30 | 11.40 ± 0.474 aB | 16.05 ± 0.201 bC | 17.18 ± 0.343 cC | 17.94 ± 0.218 dC | 18.50 ± 0.288 deC | 18.94 ± 0.288 eC | |
| 45 | 11.77 ± 0.288 aB | 11.84 ± 0.108 abB | 12.34 ± 0.108 bB | 14.92 ± 0.188 cB | 15.30 ± 0.499 cB | 16.24 ± 0.188 dB | |
| 60 | 7.94 ± 0.188 aA | 10.14 ± 0.288 bA | 10.89± 0.288 bcA | 11.52 ± 0.499 cA | 12.53 ± 0.390 dA | 14.48 ± 0.288 eA | |
| ABTS mg AAE/100g | 1 | 22.53 ± 0.492 aD | 28.98 ± 0.372 bC | 33.07 ± 0.492 cD | 33.93 ± 0.322 cC | 34.25± 0.445 cB | 37.59± 0.492 dC |
| 15 | 20.92 ± 0.492 aC | 26.73 ± 0.222 bB | 29.85 ± 0.321 cC | 30.71 ± 0.234 cB | 32.32 ± 0.322 dA | 36.19 ± 0.322 eB | |
| 30 | 19.84± 0.492 aBC | 25.44 ± 0.492 bB | 28.66± 0.492 cBC | 29.74 ±0.322 cAB | 32.53 ± 0.492 dA | 35.01± 0.492 eAB | |
| 45 | 18.55 ± 0.492 aB | 25.65 ± 0.492 bB | 28.02± 0.186 cAB | 29.41 ± 0.322 dA | 31.78 ± 0.186 eA | 34.68 ± 0.492 fA | |
| 60 | 15.33 ± 0.201 aA | 23.29 ± 0.322 bA | 27.16 ± 0.322 cA | 29.20 ± 0.186 dA | 31.03 ± 0.234 eA | 34.36 ± 0.265 fA | |
| TBARS mg MDA/Kg | 1 | 0.28 ± 0.006 fA | 0.26 ± 0.007 eA | 0.20 ± 0.001 dA | 0.11 ± 0.004 cA | 0.06 ± 0.006 bA | 0.04 ± 0.004 aA |
| 15 | 0.36 ± 0.006 eB | 0.28 ± 0.008 dA | 0.22 ± 0.011 cA | 0.13 ± 0.006 bB | 0.09 ± 0.004 aB | 0.07 ± 0.006 aB | |
| 30 | 0.42 ± 0.011 fC | 0.31 ± 0.007 eB | 0.26 ± 0.004 dB | 0.20 ± 0.008 cC | 0.17 ± 0.003 bC | 0.12 ± 0.004 aC | |
| 45 | 0.58 ± 0.009 fD | 0.49 ± 0.010 eC | 0.41 ± 0.008 dC | 0.36 ± 0.003 cD | 0.29 ± 0.003 bD | 0.20 ± 0.005 aD | |
| 60 | 0.75 ± 0.007 fE | 0.57 ± 0.005 cD | 0.48 ± 0.008 dD | 0.38 ± 0.003 cE | 0.36 ± 0.003 bE | 0.27 ± 0.005 aE |
Emulsified meat system with nanoemulsion (EMSN), 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-Azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS), and 2-thiobarbituric acid reactive substances (TBARS), Not detected (ND), gallic acid equivalents (GAE), ascorbic acid equivalents (AAE) and malonaldehyde (MDA). The lowercase letters in the superscript indicate significant differences (p < 0.05) between treatments (rows), and uppercase letters indicate significant differences in each treatment with respect to time (columns) (p < 0.05).
The results of the antioxidant activity (DPPH) assays exhibited significant differences (p < 0.05) between the treatments. The major activity was found in the treatment EMSN 5%; this activity was about 1.8-fold greater with respect to the EMSN 0% on day 60. The EMSN 0% exhibited antioxidant activity because the meat contains peptides with antioxidant properties, such as carnosine (β-alanyl-L-histidine) [35]. Sharma et al. [17] incorporated essential oils in chicken sausage and found major inhibition of DPPH radicals. The nanoemulsion contains xoconostle extracts and orange essential oil, which contain bioactive compounds, thus resulting in the increment in antioxidant activity (DPPH). The bioactive compounds inhibit free radicals [36,37,38]
The ABTS radical showed the same results as DPPH, with significant differences between the treatments (p < 0.05). Again, treatment EMSN 5% showed major antioxidant activity about 2.2-fold greater than the EMSN 0% (Table 4). Isaza et al. [31] found similar results after the incorporation of cherry extract in sausage. The phenols content and antioxidant activity were reduced with a controlled release during storage (Table 4). Again, treatments with nanoemulsions showed the best results.
Lipid oxidation showed significant differences (p < 0.05) between the treatments. Treatment EMSN 5% showed about a 2.7-fold reduction in the production of malonaldehyde (MDA) with respect to the EMSN 0%. Šojić et al. [39], Bianchin et al. [40], Erdmann et al. [41], and Ozogul et al. [42] found that the incorporation of essential oils in (their) meat systems reduced the production of malonaldehyde with respect to the control. The incorporation of the nanoemulsions with antioxidant compounds from xoconostle and orange essential oil delayed lipid oxidation, thus extending the shelf life of the EMSN.
4. Conclusions
The incorporation of the nanoemulsion in the emulsified meat system improved the nutritional contribution due to the increment in protein, inclusion of essential oils, and reduction in the loss of moisture. The texture profile showed increased hardness and chewiness. The bioactive compounds and antioxidant activities (DPPH and ABTS) incremented after the incorporation of the nanoemulsions, resulting in reduced production of malonaldehyde and minor lipid oxidation. The most favorable treatment was emulsified meat system with nanoemulsion (EMSN) 5%. Thus, the nanoemulsion extended the shelf life of the emulsified meat system.
Author Contributions
Writing—Original draft preparation I.A.-B.; Investigation A.H.-E.; Methodology R.G.-T.; Methodology N.S.-O.; Validation J.J.E.-G.; Visualization V.M.-J.; Formal analysis M.A.M.-N.; Data curation R.G.C.-M.
Funding
This research received no external funding.
Conflicts of Interest
The authors declare no conflict of interest.
References
- 1.Choi Y.-S., Choi J.-H., Han D.-J., Kim H.-Y., Lee M.-A., Kim H.-W., Jeong J.-Y., Kim C.-J. Characteristics of low-fat meat emulsion systems with pork fat replaced by vegetable oils and rice bran fiber. Meat Sci. 2009;82:266–271. doi: 10.1016/j.meatsci.2009.01.019. [DOI] [PubMed] [Google Scholar]
- 2.Šojić B., Pavlić B., Tomović V., Ikonić P., Zeković Z., Kocić-Tanackov S., Đurović S., Škaljac S., Jokanović M., Ivić M. Essential oil versus supercritical fluid extracts of winter savory (Satureja montana L.)—Assessment of the oxidative, microbiological and sensory quality of fresh pork sausages. Food Chem. 2019;287:280–286. doi: 10.1016/j.foodchem.2018.12.137. [DOI] [PubMed] [Google Scholar]
- 3.Wang X., Xie Y., Li X., Liu Y., Yan W. Effects of partial replacement of pork back fat by a camellia oil gel on certain quality characteristics of a cooked style Harbin sausage. Meat Sci. 2018;146:154–159. doi: 10.1016/j.meatsci.2018.08.011. [DOI] [PubMed] [Google Scholar]
- 4.Karovičová J., Simko P. Determination of synthetic phenolic antioxidants in food by high-performance liquid chromatography. J. Chromatogr. A. 2000;882:271–281. doi: 10.1016/S0021-9673(00)00353-8. [DOI] [PubMed] [Google Scholar]
- 5.Patel S. Plant essential oils and allied volatile fractions as multifunctional additives in meat and fish-based food products: A review. Food Addit. Contam. Part A. 2015;32:1–16. doi: 10.1080/19440049.2015.1040081. [DOI] [PubMed] [Google Scholar]
- 6.Shadman S., Hosseini S.E., Langroudi H.E., Shabani S. Evaluation of the effect of a sunflower oil-based nanoemulsion with Zataria multiflora Boiss. essential oil on the physicochemical properties of rainbow trout (Oncorhynchus mykiss) fillets during cold storage. LWT. 2017;79:511–517. doi: 10.1016/j.lwt.2016.01.073. [DOI] [Google Scholar]
- 7.Li C., Cui X., Chen Y., Liao C., Ma L.Q. Synthetic phenolic antioxidants and their major metabolites in human fingernail. Environ. Res. 2019;169:308–314. doi: 10.1016/j.envres.2018.11.020. [DOI] [PubMed] [Google Scholar]
- 8.Hernández F., Alma D., Trapala I., Angélica Gallegos V., Clemente Campos M., Rafael G., Pinedo E., José M., Guzmán M., Salvador H. Physicochemical variability and nutritional and functional characteristics of xoconostles (Opuntia spp.) accessions from México. Fruits. 2015;70:109–116. doi: 10.1051/fruits/2015002. [DOI] [Google Scholar]
- 9.Cenobio-Galindo A.D.J., Pimentel-González D.J., Del Razo-Rodríguez O.E., Medina-Pérez G., Carrillo-Inungaray M.L., Reyes-Munguía A., Campos-Montiel R.G. Antioxidant and antibacterial activities of a starch film with bioextracts microencapsulated from cactus fruits (Opuntia oligacantha) Food Sci. Biotechnol. 2019:1–9. doi: 10.1007/s10068-019-00586-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10.Espinosa-Muñoz V., RoldáN-cruz C., HernáNdez-Fuentes A., Quintero-Lira A., Almaraz-Buendía I., Campos-Montiel R. Ultrasonic-Assisted Extraction of Phenols, Flavonoids, and Biocompounds with Inhibitory Effect Against Salmonella Typhimurium and Staphylococcus Aureus from Cactus Pear. J. Food Process Eng. 2017;40:e12358. doi: 10.1111/jfpe.12358. [DOI] [Google Scholar]
- 11.Chasquibol S., Nancy Lengua C., Laura Delmás I., Rivera C.D., Bazán D., Aguirre M.R., Bravo A.M. Alimentos funcionales o fitoquímicos, clasificación e importancia. Rev. Peru. Química Ing. Química. 2003;5:9–20. [Google Scholar]
- 12.Fernández-López J., Viuda-Martos M. Introduction to the special issue: Application of essential oils in food systems. Foods. 2018;7:56. doi: 10.3390/foods7040056. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13.Shange N., Makasi T., Gouws P., Hoffman L.C. Preservation of previously frozen black wildebeest meat (Connochaetes gnou) using oregano (Oreganum vulgare) essential oil. Meat Sci. 2019;148:88–95. doi: 10.1016/j.meatsci.2018.10.012. [DOI] [PubMed] [Google Scholar]
- 14.Viuda-Martos Ruiz-Navajas Y., Fernández-López J., Pérez-Álvarez J. Antifungal activity of lemon (Citrus lemon L.), mandarin (Citrus reticulata L.), grapefruit (Citrus paradisi L.) and orange (Citrus sinensis L.) essential oils. Food Control. 2008;19:1130–1138. doi: 10.1016/j.foodcont.2007.12.003. [DOI] [Google Scholar]
- 15.Viuda-Martos Ruiz-Navajas Y., Fernández-López J., Pérez-Álvarez J.A. Effect of adding citrus fibre washing water and rosemary essential oil on the quality characteristics of a bologna sausage. LWT-Food Sci. Technol. 2010;43:958–963. doi: 10.1016/j.lwt.2010.02.003. [DOI] [Google Scholar]
- 16.Salvia-Trujillo L., Rojas-Graü A., Soliva-Fortuny R., Martín-Belloso O. Physicochemical characterization and antimicrobial activity of food-grade emulsions and nanoemulsions incorporating essential oils. Food Hydrocoll. 2015;43:547–556. doi: 10.1016/j.foodhyd.2014.07.012. [DOI] [Google Scholar]
- 17.Sharma H., Mendiratta S., Agrawal R.K., Gurunathan K., Kumar S., Singh T.P. Use of various essential oils as bio preservatives and their effect on the quality of vacuum packaged fresh chicken sausages under frozen conditions. LWT. 2017;81:118–127. doi: 10.1016/j.lwt.2017.03.048. [DOI] [Google Scholar]
- 18.Guler E., Barlas F.B., Yavuz M., Demir B., Gumus Z.P., Baspinar Y., Coşkunol H., Timur S., Başpınar Y. Bio-active nanoemulsions enriched with gold nanoparticle, marigold extracts and lipoic acid: In vitro investigations. Colloids Surf. B Biointerfaces. 2014;121:299–306. doi: 10.1016/j.colsurfb.2014.05.026. [DOI] [PubMed] [Google Scholar]
- 19.Zhang H., Tehrany E.A., Kahn C., Ponçot M., Linder M., Cleymand F. Effects of nanoliposomes based on soya, rapeseed and fish lecithins on chitosan thin films designed for tissue engineering. Carbohydr. Polym. 2012;88:618–627. doi: 10.1016/j.carbpol.2012.01.007. [DOI] [Google Scholar]
- 20.Cofrades S., Santos-López J.A., Freire M., Benedi J., Sánchez-Muniz F., Jimenez-Colmenero F. Oxidative stability of meat systems made with W1/O/W2 emulsions prepared with hydroxytyrosol and chia oil as lipid phase. LWT. 2014;59:941–947. doi: 10.1016/j.lwt.2014.06.051. [DOI] [Google Scholar]
- 21.Horwitz W. In: Official Methods of Analysis of AOAC International. Volume I, Agricultural Chemicals, Contaminants, Drugs. William H., editor. AOAC International; Gaithersburg, MD, USA: 2010. [(accessed on 24 October 2018)]. Available online: http://hdl.handle.net/10637/3158. [Google Scholar]
- 22.Cofrades S., Antoniou I., Solas M., Herrero A.M., Jimenez-Colmenero F. Preparation and impact of multiple (water-in-oil-in-water) emulsions in meat systems. Food Chem. 2013;141:338–346. doi: 10.1016/j.foodchem.2013.02.097. [DOI] [PubMed] [Google Scholar]
- 23.Singleton V.L., Orthofer R., Lamuela-Raventos R.M. [14] Analysis of Total Phenols and Other Oxidation Substrates and Antioxidants by Means of Folin-Ciocalteu Reagent. Volume 299. Elsevier BV; Amsterdam, The Netherlands: 1999. pp. 152–178. [Google Scholar]
- 24.Brand-Williams W., Cuvelier M., Berset C. Use of a free radical method to evaluate antioxidant activity. LWT. 1995;28:25–30. doi: 10.1016/S0023-6438(95)80008-5. [DOI] [Google Scholar]
- 25.Re R., Pellegrini N., Proteggente A., Pannala A., Yang M., Rice-Evans C. Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radic. Boil. Med. 1999;26:1231–1237. doi: 10.1016/S0891-5849(98)00315-3. [DOI] [PubMed] [Google Scholar]
- 26.Wang B., Pace R., Dessai A., Bovell-Benjamin A., Phillips B. Modified Extraction Method for Determining 2-Thiobarbituric Acid Values in Meat with Increased Specificity and Simplicity. J. Food Sci. 2002;67:2833–2836. doi: 10.1111/j.1365-2621.2002.tb08824.x. [DOI] [Google Scholar]
- 27.Chung C., McClements D.J. Modifying Food Texture. Elsevier; Amsterdam, The Netherlands: 2015. Structure and texture development of food-emulsion products; pp. 133–155. [Google Scholar]
- 28.Mishra P.R., Al Shaal L., Müller R.H., Keck C.M. Production and characterization of Hesperetin nanosuspensions for dermal delivery. Int. J. Pharm. 2009;371:182–189. doi: 10.1016/j.ijpharm.2008.12.030. [DOI] [PubMed] [Google Scholar]
- 29.Gago C.M.L., Artiga-Artigas M., Antunes M.D.C., Faleiro M.L., Miguel M.G., Martín-Belloso O. Effectiveness of nanoemulsions of clove and lemongrass essential oils and their major components against Escherichia coli and Botrytis cinerea. J. Food Sci. Technol. 2019;56:2721–2736. doi: 10.1007/s13197-019-03762-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 30.Donsi’ F., Annunziata M., Sessa M., Ferrari G. Nanoencapsulation of essential oils to enhance their antimicrobial activity in foods. LWT. 2011;44:1908–1914. doi: 10.1016/j.lwt.2011.03.003. [DOI] [Google Scholar]
- 31.Ushikubo F., Cunha R., Cunha R. Stability mechanisms of liquid water-in-oil emulsions. Food Hydrocoll. 2014;34:145–153. doi: 10.1016/j.foodhyd.2012.11.016. [DOI] [Google Scholar]
- 32.Bolger Z., Brunton N.P., Monahan F.J. Impact of inclusion of flaxseed oil (pre-emulsified or encapsulated) on the physical characteristics of chicken sausages. J. Food Eng. 2018;230:39–48. doi: 10.1016/j.jfoodeng.2018.02.026. [DOI] [Google Scholar]
- 33.Youssef M., Barbut S., Youssef M. Effects of protein level and fat/oil on emulsion stability, texture, microstructure and color of meat batters. Meat Sci. 2009;82:228–233. doi: 10.1016/j.meatsci.2009.01.015. [DOI] [PubMed] [Google Scholar]
- 34.Isaza Y.L., Restrepo D.A., López J.H., Ochoa O.A., Cabrera K.R. Evolution of the antioxidant capacity of frankfurter sausage model systems with added cherry extract (Prunus Avium L.) during refrigerated storage. Vitae. 2011;18:251–260. [Google Scholar]
- 35.Weiss J., Gibis M., Schuh V., Salminen H. Advances in ingredient and processing systems for meat and meat products. Meat Sci. 2010;86:196–213. doi: 10.1016/j.meatsci.2010.05.008. [DOI] [PubMed] [Google Scholar]
- 36.Kuskoski E.M., Asuero A.G., Troncoso A.M., Mancini-Filho J., Fett R. Aplicación de diversos métodos químicos para determinar actividad antioxidante en pulpa de frutos. Food Sci. Technol. 2005;25:726–732. doi: 10.1590/S0101-20612005000400016. [DOI] [Google Scholar]
- 37.Munguía A.R., Nieto E.A., Beristain C., Sosa F.C., Carter E.V. Propiedades antioxidantes del maguey morado (Rhoeo discolor) CyTA J. Food. 2009;7:209–216. doi: 10.1080/19476330903010177. [DOI] [Google Scholar]
- 38.Kumar Y., Yadav D.N., Ahmad T., Narsaiah K. Recent Trends in the Use of Natural Antioxidants for Meat and Meat Products. Compr. Rev. Food Sci. Food Saf. 2015;14:796–812. doi: 10.1111/1541-4337.12156. [DOI] [Google Scholar]
- 39.Šojić B., Tomović V., Kocic-Tanackov S., Skaljac S., Ikonic P., Džinić N., Živković N., Jokanović M., Tasic T., Kravić S. Effect of nutmeg (Myristica fragrans) essential oil on the oxidative and microbial stability of cooked sausage during refrigerated storage. Food Control. 2015;54:282–286. doi: 10.1016/j.foodcont.2015.02.007. [DOI] [Google Scholar]
- 40.Bianchin M., Pereira D., Dos Reis A.S., Almeida J.D.F., Da Silva L.D., De Moura C., Carpes S.T. Rosemary Essential Oil and Lyophilized Extract as Natural Antioxidant Source to Prevent Lipid Oxidation in Pork Sausage. Adv. J. Food Sci. Technol. 2017;13:210–217. doi: 10.19026/ajfst.13.5070. [DOI] [Google Scholar]
- 41.Erdmann M.E., Lautenschlaeger R., Zeeb B., Gibis M., Weiss J. Effect of differently sized O/W emulsions loaded with rosemary extract on lipid oxidation in cooked emulsion-type sausages rich in n-3 fatty acids. LWT. 2017;79:496–502. doi: 10.1016/j.lwt.2016.03.022. [DOI] [Google Scholar]
- 42.Ozogul Y., Yuvka İ., Ucar Y., Durmus M., Kösker A.R., Öz M., Ozogul F. Evaluation of effects of nanoemulsion based on herb essential oils (rosemary, laurel, thyme and sage) on sensory, chemical and microbiological quality of rainbow trout (Oncorhynchus mykiss) fillets during ice storage. LWT-Food Sci. Technol. 2017;75:677–684. doi: 10.1016/j.lwt.2016.10.009. [DOI] [Google Scholar]