provenance: AgriculturalAndFoodSciences,2022-2022-Agricultural And Food Sciences-2022.jsonl.gz.gz

text:
Supercritical Fluid Extraction of Fucoxanthin from the Diatom Phaeodactylum tricornutum and Biogas Production through Anaerobic Digestion

Phaeodactylum tricornutum is the marine diatom best known for high-value compounds that are useful in aquaculture and food area. In this study, fucoxanthin was first extracted from the diatom using supercritical fluid extraction (SFE) and then using the extracted diatom-like substrate to produce bioenergy through anaerobic digestion (AD) processes. Factors such as temperature (30 °C and 50 °C), pressure (20, 30, and 40 MPa), and ethanol (co-solvent concentration from 10% to 50% v/v) were optimized for improving the yield, purity, and recovery of fucoxanthin extracted using SFE. The highest yield (24.41% w/w) was obtained at 30 MPa, 30 °C, and 30% ethanol but the highest fucoxanthin purity and recovery (85.03mg/g extract and 66.60% w/w, respectively) were obtained at 30 MPa, 30 °C, and 40%ethanol. Furthermore, ethanol as a factor had the most significant effect on the overall process of SFE. Subsequently, P.tricornutum biomass and SFE-extracted diatom were used as substrates for biogas production through AD. The effect of fucoxanthin was studied on the yield of AD, which resulted in 77.15 ± 3.85 LSTP CH4/kg volatile solids (VS) and 56.66 ± 1.90 LSTP CH4/kg VS for the whole diatom and the extracted P.tricornutum, respectively. Therefore, P.tricornutuman can be considered a potential source of fucoxanthin and methane and both productions will contribute to the sustainability of the algae-biorefinery processes.

Introduction
Diatoms are unicellular photosynthetic eukaryotes that are commonly found in marine ecosystems and moist terrestrial habitats. They contribute to about 20-25% of global carbon dioxide fixation through photosynthesis and play an important role in the global silicon cycle [1,2]. In addition, they are a part of the base of marine trophic networks worldwide and contribute to at least 20% of annual primary productivity, which is equivalent to tropical forests [3,4]. Among the organic molecules produced by diatoms, fatty acids or avoids all the problems associated with the use of inhibitory organic solvents by facilitating the valorization of extracted microalgae through AD.
In this study, we performed supercritical carbon dioxide (SCO 2 ) extraction of fucoxanthin from the diatom P. tricornutum and AD of the extracted microalga under microalgae biorefinery frame. We also studied the effects of factors such as temperature, pressure, and co-solvent concentration on variables such as the total extraction yield, fucoxanthin purity, and recovery. Finally, the biochemical composition of P. tricornutum and methane yields before and after the SFE process were studied to determine how the use of a green extraction system affects methane production. In this study, we performed supercritical carbon dioxide (SCO2) extraction of fucoxanthin from the diatom P. tricornutum and AD of the extracted microalga under microalgae biorefinery frame. We also studied the effects of factors such as temperature, pressure, and co-solvent concentration on variables such as the total extraction yield, fucoxanthin purity, and recovery. Finally, the biochemical composition of P. tricornutum and methane yields before and after the SFE process were studied to determine how the use of a green extraction system affects methane production.

Effects of Pressure and Temperature on the Yield of Fucoxanthin from P. tricornutum
Total extraction yields for each operating conditions (temperature of 30 °C and 50 °C and pressure of 20, 30, and 40 MPa) are summarized in Figure 1. The results are expressed as the percentage of g extract per g dry weight of P. tricornutum, and the values were obtained at the end of extraction (60 min). This extraction time was established as optimal in previous studies on carotenoids from microalgae [30] and was considered profitable for minimizing its operational costs in the process. In general, total extraction yields were slightly affected by the combination of pressure and temperature factors. The highest total yields were without significant differences (p < 0.05) between them, if arranged from the highest to the lowest value, at the highest pressure and temperature (40 MPa at 50 °C, 3.14 ± 0.03%), followed by 30 MPa at 30 °C and 30 MPa at 50 °C (3.05 ± 0.05% and 3.00 ± 0.09%), respectively. On the other hand, the lowest extraction yields, which in turn did not have significant differences (p < 0.05) between them, were obtained at 20 MPa at 30 °C (2.55% w/w) together with the condition of 40 MPa at 30 °C and 20 MPa at 50 °C, which were 2.61 and 2.73% w/w, respectively ( Figure 1).
As shown in Table 1   The results are expressed as the percentage of g extract per g dry weight of P. tricornutum, and the values were obtained at the end of extraction (60 min). This extraction time was established as optimal in previous studies on carotenoids from microalgae [30] and was considered profitable for minimizing its operational costs in the process. In general, total extraction yields were slightly affected by the combination of pressure and temperature factors. The highest total yields were without significant differences (p < 0.05) between them, if arranged from the highest to the lowest value, at the highest pressure and temperature (40 MPa at 50 • C, 3.14 ± 0.03%), followed by 30 MPa at 30 • C and 30 MPa at 50 • C (3.05 ± 0.05% and 3.00 ± 0.09%), respectively. On the other hand, the lowest extraction yields, which in turn did not have significant differences (p < 0.05) between them, were obtained at 20 MPa at 30 • C (2.55% w/w) together with the condition of 40 MPa at 30 • C and 20 MPa at 50 • C, which were 2.61 and 2.73% w/w, respectively ( Figure 1).
As shown in Table 1 and the operating conditions, increasing the pressure at constant temperature increases the CO 2 density from 0.890 to 0.988 g/mL for pressures of 20-40 MPa at 30 • C and from 0.784 to 0.923 g/mL for pressures of 20-40 MPa at 50 • C. Under these conditions of increased pressure, the total extraction yield ( Figure 1) was slightly improved without significant differences (p < 0.05) for 30  and it could be due to the volatility of the extracts under these SFE conditions, or the component solubility is lowered as the polarity and/or the molecular weights of the solutes are increased [19].  Table 1 also shows the purity, calculated as mg fucoxanthin extracted by SFE per g of total extract, and recovery, calculated as the percentage of the weight of fucoxanthin extracted by conventional extraction per grams of biomass as the reference weight of this pigment (equal to 9.82 ± 0.70 mg/g biomass) from of P. tricornutum. The different temperature and pressure conditions had a significant effect on these fucoxanthin variables, according to Tukey's test. Particularly, the highest fucoxanthin purity and recovery were reached for a pressure of 30 MPa at 30 • C (60.62 ± 0.40 mg/g and 18.82 ± 0.12% w/w, respectively), being significant (p < 0.05) for the rest of the processes. Under these conditions, the density and superficial CO 2 velocity were 0.948 g/mL and 0.401 mm/s, respectively, being the second-highest density and lowest superficial velocity, exceeded only by 30 • C and 40 MPa, whose values were 0.988 g/mL and 0.384 mm/s. However, in this treatment, the purity and recovery of fucoxanthin were not the highest. As shown in Table 1, the values of the superficial CO 2 velocities were in the range of 0.384-0.484 mm/s, indicating slight variation, because the mass flow rate (3.62 g CO 2 /min) and the area of the extraction vessel (1589 cm 2 ) were constant, the variations were only due to the density of CO 2 that depends on temperature and pressure.
At low temperatures, the purity was higher than that at high temperatures under the same pressure values. This tendency was similar for fucoxanthin recoveries except for 40 MPa and 50 • C, which was approximately two-fold higher than that at 40 MPa and 30 • C. In contrast, at low pressure and under both temperature conditions, fucoxanthin purity and recovery were the lowest.
The particle size distribution is another important parameter in SFE as it directly affects the porosity and physical properties of the extraction bed. The particle diameter is inversely proportional to the total extraction yield [31,32]. On the other hand, a reduction in the final particle diameter increases the surface-to-volume ratio of the substrate [33]. The mean particle size of the Phaeodactylum tricornutum powder was D P = 0.074 ± 0.004 mm, which is considered an ultrafine powder. Crampon et al. [34] reported that the smaller the particle, the faster the kinetics of the extraction, thus higher the yields. The disintegration of cells is essential in the recovery of intracellular products from algae.
The effect of temperature on extraction yield is more complex than that of pressure. This behavior is known as "crossover pressure" and is defined as the point where the vapor pressure of the compounds in the extract is more pronounced than the effect of solvent density over the solubility of the compounds [35,36]. The crossover region is a phenomenological observation that is supported by a relevant number of experimental studies. This phenomenon refers to the regions of temperature and pressure in the near-critical fluid state where the solute solubility decreases with an isobaric increase in temperature [37].
The effect of temperature on solubility is complex, and the crossover of the solubility isotherm occurs when pressure is considered as a variable. This phenomenon is affected by solvent density and solute vapor pressure. The vapor pressure increases when temperature and solubility increase; on the contrary, density and solvent power decrease. Consequently, solubility decreases because the density of the solvent is the dominant factor. After passing the crossover area, and when the temperature is increased, the vapor pressure of the solute becomes a dominant factor, and the solubility increases. However, this contrasting behavior is often attributed to solvent density and solute vapor pressure [38].
Foster et al. [39] reported that crossover pressure is a phenomenological observation that appears to reflect a characteristic of the solute-solvent system. Furthermore, it is also of fundamental significance in providing a direct indicator of the reliability and consistency of experimental solubility data. As a consequence, it may also be used to introduce a phenomenological constraint on thermodynamic models proposed to describe solutesupercritical fluid phase equilibria. This means that, at pressures lower than the crossover pressure, the effect of CO 2 density is more relevant, whereas for pressures higher than the crossover pressure, the effect of vapor pressure at the determined temperature is more relevant for increasing the extraction yield. This is a common behavior for carotenoids obtained by SFE from vegetable matrices [37].
Fabrowska et al. [40] extracted carotenoids and phenolic compounds from the freshwater macroalgae Cladophora glomerata by optimization of the same parameters included in our study (temperature, pressure, and % ethanol as a co-solvent). The extraction yields were less than our yields without the use of extractants such as 0.6-0.9% w/w. The higher value of carotenoids recovery, represented with fucoxanthin as a majority, was 6.17 ± 0.24 mg/g of the extract under~30 MPa and 50 • C, being similar conditions, except for the temperature. Ruiz-Domínguez et al. [41] used a Box-Behnken design with desirability function to determine the bioactive composition from the microalga Isochrysis galbana by SFE, and the parameters studied were pressure , temperature (40-60 • C), and co-solvent (0-8% ethanol) with a CO 2 flow rate of 7.2 g/min for 120 min. The extraction yield without the presence of ethanol was in a similar range (from 1.09% to 2.28% w/w), and the highest was 2.28 ± 0.11% w/w under 40 MPa at 50 • C. They also determined total carotenoids recovery (included fucoxanthin) with an optimum value of 16.61 ± 0.74% at 40 MPa and 50 • C without co-solvent and 77.93 ± 2.87% at 30 MPa, 60 • C, and 8% v/v ethanol. Roh et al. [42] extracted fucoxanthin and polyphenols from the brown macroalga Undaria pinnatifida using SCO 2 and ethanol as co-solvent. The highest yields of fucoxanthin in the presence of ethanol were at~20 MPa at~50 • C with a value of fucoxanthin recovery much lower than our results (approximately 0.00753 µg fucoxanthin per g freeze-dried sample). Therefore, our results showed that an increase in pressure (from 20 to 30 MPa) at a constant temperature (30 • C) increased the solvating power of carbon dioxide as supercritical fluids favoring fucoxanthin extraction. However, the temperature is a sensitive factor at high ranges, as it can degrade the targeted compound even though it can increase the solubility of this component [43].
Although all results of yield and recovery of fucoxanthin might be improved, the optimal conditions of pressure and temperature for the highest purity and recovery of fucoxanthin were 30 MPa and 30 • C and were selected for evaluating the behavior under ethanol as an extractant by SFE.

Effects of Ethanol as a Co-Solvent Factor on the Yield of Fucoxanthin from Phaeodactylum tricornutum
Many studies have reported that the use of ethanol as an extractant improves the yields of fucoxanthin due to its polarity and affinity with carotenoid molecules [44,45]. The optimal pressure and temperature for the highest purity and recovery of fucoxanthin along with the presence of co-solvent were used for evaluating the effect on the extraction process. Figure 2 shows the total extraction yield obtained under different concentrations of ethanol from 10% to 50% v/v. Here, the co-solvent improved the extraction yield, being at 30% v/v of ethanol, the highest value was obtained with 24.41% w/w, followed by 20% v/v of co-solvent with a value of 14.28% w/w. However, the lowest value was obtained at 10% and 50% v/v of ethanol, with 5.65% and 5.18% w/w of total extraction yields, respectively.
Mar. Drugs 2022, 20, x 6 of 20 along with the presence of co-solvent were used for evaluating the effect on the extraction process. Figure 2 shows the total extraction yield obtained under different concentrations of ethanol from 10% to 50% v/v. Here, the co-solvent improved the extraction yield, being at 30% v/v of ethanol, the highest value was obtained with 24.41% w/w, followed by 20% v/v of co-solvent with a value of 14.28% w/w. However, the lowest value was obtained at 10% and 50% v/v of ethanol, with 5.65% and 5.18% w/w of total extraction yields, respectively. In general, the effect of co-solvent was significant; 30% v/v ethanol improved total yield to 8-fold higher than that using 30 MPa, 30 °C, and only carbon dioxide as a solvent. Many studies have reported that SFE is an excellent technique for the selective extraction of bioactive compounds such as carotenoids from various microalgae [16,43,46]. As a general rule, the extraction improves when SCO2 is used, whereas the presence of co-solvent or extractant such as ethanol enhances the efficiency of the supercritical process but disparages the selectivity [16].
On the other hand, Table 2 shows fucoxanthin purity and recovery under 30 MPa and 30 °C at different percentages of co-solvent. Different superscripted alphabets (a-c) indicate significant differences (p < 0.05) and the data are expressed as mean ± standard deviation (n = 3, SD ≤ 5%).
The highest values with no significant differences in fucoxanthin purity were 85.03 ± 7.67 mg/g and 74.73 ± 2.45 mg/g extract using 40 and 50% v/v of ethanol, respectively. In general, the effect of co-solvent was significant; 30% v/v ethanol improved total yield to 8-fold higher than that using 30 MPa, 30 • C, and only carbon dioxide as a solvent. Many studies have reported that SFE is an excellent technique for the selective extraction of bioactive compounds such as carotenoids from various microalgae [16,43,46]. As a general rule, the extraction improves when SCO 2 is used, whereas the presence of co-solvent or extractant such as ethanol enhances the efficiency of the supercritical process but disparages the selectivity [16].
On the other hand, Table 2 shows fucoxanthin purity and recovery under 30 MPa and 30 • C at different percentages of co-solvent. Different superscripted alphabets (a-c) indicate significant differences (p < 0.05) and the data are expressed as mean ± standard deviation (n = 3, SD ≤ 5%).
The highest values with no significant differences in fucoxanthin purity were 85.03 ± 7.67 mg/g and 74.73 ± 2.45 mg/g extract using 40 and 50% v/v of ethanol, respectively. However, the lowest purity was 13.75 ± 0.65 mg/g extract at 30% v/v of co-solvent despite the extraction yield being higher in this condition.
The highest fucoxanthin recovery with significant differences among others was 66.60 ± 6.00% w/w under 40% v/v of ethanol, whereas the lowest was 12.69 ± 1.14% w/w using 10% v/v of ethanol. The fucoxanthin purity and recovery reached the maximum value under the same conditions of 30 MPa, 30 • C, and 40% v/v ethanol. As reported by Gómez-Loredo et al. [47], an increasing concentration of co-solvent could induce a decrease in the purity and recovery of fucoxanthin as other compounds that were the part of the extract could transfer with this carotenoid. At times, up to a certain value, the addition of co-solvent has no further effect. This could happen at 50% v/v ethanol where fucoxanthin purity and recovery begin to decrease.
Extraction of fucoxanthin from different natural sources using conventional solvent, pressurized liquid extraction, or supercritical fluids extraction has been reported. For example, Conde et al. [48] showed ethanol concentration (0.5-10% v/v) as a significant factor in the supercritical CO 2 extraction process from the macroalgae Sargassum muticum. The improvement in ethanol concentration resulted in total yields up to 3 times higher, radical scavenging capacity up to 2.5 times higher, and fucoxanthin extraction yield up to 90 times higher, thus reaching approximately 12 mg fucoxanthin/100 mg dry biomass under 20 MPa at 30 • C and 10% v/v ethanol. Gilbert-López et al. [49] compared MAE and PLE as green techniques for the extraction of bioactive compounds from P. tricornutum. The total yield (23.95% w/w), fucoxanthin purity, and its recovery (32.29 mg/g extract and 54.41% w/w, respectively) were the highest under 50 • C and 100% ethanol using PLE. Our results of fucoxanthin purity and recovery are higher than these results, although the P. tricornutum strain described by Gilbert-López et al. [49] reached up to 1.5-times higher concentration of fucoxanthin by conventional extraction than our results (maceration in acetone during 24 h at room temperature was around of 59.5 ± 4.7 mg/g extract). All results corroborated that P. tricornutum is an important source of bioactive compounds such as fucoxanthin and is better than other species such as macroalgae. Table 3 presents the physicochemical characterization of the diatom P. tricornutum and supercritical-fluid-extracted P. tricornutum. As shown, most of the analyzed parameters have maintained similar values in both groups. Only metal content, which increased in all cases after SFE, and the P T -P 2 O 5 parameter decreased by four times after SFE. It should be highlighted that the metal content reached non-toxic or inhibiting concentrations. The maintenance of nutrients such as nitrogen (N-TKN) or phosphorous (P T -P 2 O 5 ) in the diatom P. tricornutum is essential for the microorganisms responsible for performing AD.

Biochemical Composition and the Effects of SFE on AD Using P. tricornutum
The comparison of the biochemical composition of P. tricornutum biomass and other strains is presented in Table 4. Biogas production from microalgae depends on their biochemical composition and other factors such as operating conditions, cell wall nature, C/N ratios and ammonia release, and sodium in marine species [50][51][52]. The analysis of the biochemical composition may help in the comprehension of the methane yields obtained. The best theoretical methane yields obtained are for lipids, followed by proteins and carbohydrates, with values 1.014 L CH 4 /g VS, 0.851 L CH 4 /g VS, and 0.415 L CH 4 /g VS, respectively [51]. These values could be accurate considering the composition of the specific microalga species.
Generally, the exhausted diatom has the lowest macronutrient content due to the extraction process, highlighting the decrease in lipid and protein content by more than half. The biochemical composition of pretreated P. tricornutum markedly decreased with the extraction of 68.99% and 60.19% of lipids and proteins, respectively, until the values of 5.01 ± 0.01% and 14.41 ± 0.62% w/w, respectively, were obtained. However, only 29.15% of carbohydrates were extracted. On the other hand, the protein content differed significantly (p ≤ 0.05) among the strains, followed by lipid and carbohydrate contents. The protein content ranged from 38.8 ± 0.11% to 26.95 ± 0.05% w/w (excluding P. tricornutum after SFE) with P. tricornutum F&M-M4 showing the highest value (Table 4). For carbohydrate content, all strains showed similar values with slight differences. Particularly, the carbohydrate content of P. tricornutum in our study was higher (18.80 ± 1.12% w/w) than the P. tricornutum carbohydrate content (16.91 ± 1.61% w/w) reported by Di Lena et al. [53]. P. tricornutum F&M-M4 had the highest lipid content, followed by a diatom described by Bernaerts et al. [54] (17.1 ± 0.9% w/w). In the case of extracted P. tricornutum, the lipid content was significantly decreased compared with the initial content in the biomass before extraction because especially the lipid fraction was recovered through supercritical CO 2 -extraction technology.
Globally, such results have confirmed P. tricornutum as complete and nutritive biomass; hence, it is important in aquaculture feeds and food applications [5,9,10,55]. Nevertheless, the biochemical compositions of P. tricornutum found in previous studies are variable and highly dependent on strains, culturing conditions, and growth stages [54,56]. Table 4. Biochemical composition of the diatom P. tricornutum before and after supercritical fluid extraction compared with the composition reported in previous studies.

Strain/Diatom
Proteins (% w/w) * Sum of storage polysaccharides (SPS) and cell wall polysaccharides (CWPS). Note: Different superscripted alphabets (a-e) indicate significant differences (p < 0.05) and the data are expressed as mean ± standard deviation (n = 3, SD ≤ 5%). Nucleic acids and ash account for the percentage left to make 100% wt of proximal composition.

Lipids
As shown in Table 4, our supercritical-fluid-extracted P. tricornutum or exhausted biomass was rich in proteins, carbohydrates, and to a lesser extent in lipids. This could be used in different integrated processes in algae biorefinery. Hence, in our study, the production of bioenergy in the form of biogas was evaluated.

BMP of Diatom P. tricornutum and Extracted P. tricornutum by SFE
Typically, the yield of biogas obtained by AD is associated with the biochemical composition and VS content of the biomass used as a substrate. Particularly, the AD of microalgae can be affected by different factors such as low C/N ratio, thick cell walls banking on the microalgal genus, and high concentrations of sodium for marine species [50,52,58].
P. tricornutum is the only species in the genus Phaeodactylum, which is rich in bioactive compounds such as fucoxanthin and fatty acids [10,55]. However, this genus has a silicious wall, one of the main drawbacks when considering the microalgae for AD that could affect the results of this process. When intracellular compounds of interest such as fucoxanthin are extracted by SFE from microalgae, the action of temperature, pressure, and the use of ethanol as a co-solvent can be considered as a pretreatment. SFE helps in the extraction of fucoxanthin, but the breakage of cell structures also favors the release of other compounds, which facilitates the action of microorganisms that perform the hydrolytic steps of AD. After SFE, the cell wall and structure breaks to some extent, and the remaining compounds and cell material are released into the surrounding environment, which can be used as substrates to feed anaerobic microorganisms [59].
The methane yields obtained for the whole P. tricornutum and the extracted diatom were 77.2 ± 3.85 LSTP CH 4 /kg VS and 56.7 ± 1.90 LSTP CH 4 /kg VS, respectively (Figure 3). algae can be affected by different factors such as low C/N ratio, thick cell walls banking on the microalgal genus, and high concentrations of sodium for marine species [50,52,58].
P. tricornutum is the only species in the genus Phaeodactylum, which is rich in bioactive compounds such as fucoxanthin and fatty acids [10,55]. However, this genus has a silicious wall, one of the main drawbacks when considering the microalgae for AD that could affect the results of this process. When intracellular compounds of interest such as fucoxanthin are extracted by SFE from microalgae, the action of temperature, pressure, and the use of ethanol as a co-solvent can be considered as a pretreatment. SFE helps in the extraction of fucoxanthin, but the breakage of cell structures also favors the release of other compounds, which facilitates the action of microorganisms that perform the hydrolytic steps of AD. After SFE, the cell wall and structure breaks to some extent, and the remaining compounds and cell material are released into the surrounding environment, which can be used as substrates to feed anaerobic microorganisms [59].
The methane yields obtained for the whole P. tricornutum and the extracted diatom were 77.2 ± 3.85 LSTP CH4/kg VS and 56.7 ± 1.90 LSTP CH4/kg VS, respectively (Figure 3). In both cases, a methane yield of around 55 LSTP CH4/kg VS was obtained in less than 10 h, highlighting the fast biodegradability of the substrate and microbial activity at the beginning of AD. Caporgno et al. [60] obtained methane yields of 257 ± 8 mL of CH4/g VS and 180 ± 6 mL of CH4/g VS, respectively, for P. tricornutum after lipid extraction for biodiesel production with traditional solvents such as hexane and methanol/hexane mixture. Zhao et al. [26] obtained the yield of 339 ± 13 mL of CH4/g VS for the lipid-extracted microalgal biomass using hexane and evaporating it after the extraction at 105 °C in a hood. Studies on the AD of P. tricornutum are limited, and the methane yields reported by Caporgno et al. In both cases, a methane yield of around 55 LSTP CH 4 /kg VS was obtained in less than 10 h, highlighting the fast biodegradability of the substrate and microbial activity at the beginning of AD. Caporgno et al. [60] obtained methane yields of 257 ± 8 mL of CH 4 /g VS and 180 ± 6 mL of CH 4 /g VS, respectively, for P. tricornutum after lipid extraction for biodiesel production with traditional solvents such as hexane and methanol/hexane mixture. Zhao et al. [26] obtained the yield of 339 ± 13 mL of CH 4 /g VS for the lipid-extracted microalgal biomass using hexane and evaporating it after the extraction at 105 • C in a hood. Studies on the AD of P. tricornutum are limited, and the methane yields reported by Caporgno et al. [60], Zhao et al. [26], Frigon et al. [61], or Zamalloa et al. [62] could not be compared with the yields obtained in the present study because the extraction methods were not the same, nor did the extracted compounds include fucoxanthin, and the extraction of different compounds can affect AD methane yields differently.
In addition, a high or low concentration of a co-solvent (as a remainder) at the end of the extraction process of added high-value products can increase methane production when they are biodegradable. In our study, the methane yield obtained for the whole diatom was higher (77.2 ± 3.85 LSTP CH 4 /kg VS) than that obtained for the extracted diatom (56.7 ± 1.90 LSTP CH 4 /kg VS), resulting in a 26.6% reduction in the methane yield obtained from the whole microorganism. However, from the environmental and economic point of view, through the AD of the extracted P. tricornutum, biogas was obtained, which could reduce the energy costs for fucoxanthin extraction by SFE and/or contribute to the reduction in the costs of growing new microalgal biomass conferring the circularity to the process. Table 5 shows the characterization of the effluents or digests obtained after BMP assays of the microalgae P. tricornutum and the supercritical-fluid-extracted P. tricornutum. Conductivity was stable in both microalgae, with values around 17.75 to 18.14 mS/cm in all effluents obtained and loads assayed. The pH in these effluents increased with the second load in the BMP assays of both diatoms. The pH values obtained in the second load of the supercritical-fluid-extracted P. tricornutum were 9.05 ± 0.02, which are especially high for an anaerobic system, for which the optimal pH range is from 6.7 to 7.5, close to neutral ranges. In anaerobic processes and at this pH, VFA and ammonia almost dissociate.   Figure 4 shows the values of T-Alk measured in the effluents obtained after the BMP assays of diatom P. tricornutum and supercritical-fluid-extracted P. tricornutum.

Characteristics of Effluents after AD
T-Alk is an important parameter for AD systems. T-Alk gives anaerobic systems the capacity to cope with pH fluctuations. Most anaerobic reactors have T-Alks between 1500 and 5000 mg CaCO 3 /L, although values above 5000 mg CaCO 3 /L found in some reactors exhibited a slightly negative effect on the process [63]. In the present study, T-Alk values of the effluents were high, 9678 ± 90 mg CaCO 3 /L and 10,098 ± 105 mg CaCO 3 /L for the first and the second loads assayed with P. tricornutum and 9815 ± 115 and 10,300 ± 95 mg CaCO 3 /L for the first and second loads with supercritical-fluid-extracted P. tricornutum, respectively. This increase in T-Alk could be responsible for the increase in ammonia concentration. The initial growth of ammonia in the media can increase buffer capacity [64]; however, the high concentrations of ammonia can inhibit methanogenesis. solids, VS: volatile solids, N-NH4+: ammoniacal nitrogen, TKN: total Kjeldahl Nitrogen, DL: Detection Limit, Phaeodactylum Tricornutum (P. tricornutum). Figure 4 shows the values of T-Alk measured in the effluents obtained after the BMP assays of diatom P. tricornutum and supercritical-fluid-extracted P. tricornutum. T-Alk is an important parameter for AD systems. T-Alk gives anaerobic systems the capacity to cope with pH fluctuations. Most anaerobic reactors have T-Alks between 1500 and 5000 mg CaCO3/L, although values above 5000 mg CaCO3/L found in some reactors exhibited a slightly negative effect on the process [63]. In the present study, T-Alk values of the effluents were high, 9678 ± 90 mg CaCO3/L and 10,098 ± 105 mg CaCO3/L for the first and the second loads assayed with P. tricornutum and 9815 ± 115 and 10,300 ± 95 mg CaCO3/L for the first and second loads with supercritical-fluid-extracted P. tricornutum, respectively. This increase in T-Alk could be responsible for the increase in ammonia concentration. The initial growth of ammonia in the media can increase buffer capacity [64]; however, the high concentrations of ammonia can inhibit methanogenesis.
In anaerobic digesters, ammonium and free ammonia are in equilibrium, and free ammonia, a form of nitrogen, is more dangerous for these systems. Free ammonia concentrations are usually higher at higher pH and temperatures, reaching an inhibitory concentration of 150 mg N-NH3/L [65]. In the present study, N-NH4 + concentration increased from 1472 ± 60 to 1653 ± 50 mg/L when moving from the first load to the second load in the reactors digesting P. tricornutum. On the other hand, N-NH4 + concentrations were almost constant for the first and second loads of the extracted P. tricornutum, with values of 1265 ± 5 and 1290 ± 10 mg/L for the first and second load assayed, respectively ( Figure  5A). In anaerobic digesters, ammonium and free ammonia are in equilibrium, and free ammonia, a form of nitrogen, is more dangerous for these systems. Free ammonia concentrations are usually higher at higher pH and temperatures, reaching an inhibitory concentration of 150 mg N-NH 3 /L [65]. In the present study, N-NH 4 + concentration increased from 1472 ± 60 to 1653 ± 50 mg/L when moving from the first load to the second load in the reactors digesting P. tricornutum. On the other hand, N-NH 4 + concentrations were almost constant for the first and second loads of the extracted P. tricornutum, with values of 1265 ± 5 and 1290 ± 10 mg/L for the first and second load assayed, respectively ( Figure 5A). The values obtained for the whole diatom P. tricornutum were high and close to inhibitory values obtained in anaerobic environments, between 1500 and 3000 mg N-NH4 + /L [66]. Inhibitory concentration values can be higher in an adapted process; some studies have shown the tolerance of microalgae to ammonia concentrations between 1500-7000 mg N-NH4 + /L [67]. Factors such as the origin of the inoculum, temperature, and types of substrates may affect this wide range of inhibitory concentrations of N-NH4 + [65].
The N-TKN ( Figure 5B) present in the effluents obtained after the first and second load of the diatom P. tricornutum were higher than that obtained after the first and second load of the supercritical-fluid-extracted P. tricornutum, obtaining values of 2013 ± 25 mg/L The values obtained for the whole diatom P. tricornutum were high and close to inhibitory values obtained in anaerobic environments, between 1500 and 3000 mg N-NH 4 + /L [66]. Inhibitory concentration values can be higher in an adapted process; some studies have shown the tolerance of microalgae to ammonia concentrations between 1500-7000 mg N-NH 4 + /L [67].
Factors such as the origin of the inoculum, temperature, and types of substrates may affect this wide range of inhibitory concentrations of N-NH 4 + [65]. The N-TKN ( Figure 5B) present in the effluents obtained after the first and second load of the diatom P. tricornutum were higher than that obtained after the first and second load of the supercritical-fluid-extracted P. tricornutum, obtaining values of 2013 ± 25 mg/L and 2169 ± 25 mg/L for the first and second loads of the whole P. tricornutum, respectively. These values were lower for the extracted P. tricornutum, 1720 ± 10 and 1812 ± 15 mg/L for the first and second loads, respectively. These values could be responsible for a 39.8% reduction in proteins, which decreased from 36.20% to 14.41% (w/w) for the diatom P. tricornutum due to SFE, as described in Section 2.3 and Table 4. Figure 6 shows the VFA evolution in each load studied for the diatom P. tricornutum and supercritical-fluid-extracted P. tricornutum. VFAs in the effluents after the BMP assays showed a decreasing trend at the end of the second load, decreasing values from 195 ± 5 mg C/L to 87 ± 5 mg C/L for the non-extracted diatom and from 190 ± 10 mg C/L to 154 ± 5 mg C/L for the supercritical-fluid-extracted P. tricornutum. The decrease in acidity was an indicator of the presence of an adequate amount of methanogenic bacteria. Finally, it seems that metal content provided by the own diatom does not apparently affect the AD process. The use of microalgae (including diatoms) in co-digestion with rich carbon substrates is common for obtaining adequate nutrients and metal balance for the anaerobic system [68].

Biomass and Chemicals
The diatom Phaeodactylum tricornutum (CIB-41 from CIBNOR (https://www.cibnor.gob.mx/) La Paz, Mexico) was obtained from the Mexican Company "Microalgas Oleas de México S.A." (Guadalajara, Mexico). The diatom was cultured in F/2 culture medium as reported previously by Guillard [69] and harvested at the end of the exponential phase. The microalga (diatom) was provided freeze-dried (freeze-dry system Labconco Freezone 18 L Benchtop Freeze Dry System, Kansas, MO, USA), packed in vacuum-sealed plastic bags and then was stored at 4 ± 2 °C in the dark until use. All solutions used in this study were of analytical grade except for the standard solutions and chromatographic solvents. For supercritical fluid extraction, carbon dioxide and ethanol were purchased from Indura Air Products Group (Santiago, Chile) and Sigma-Aldrich (Santiago, Chile), respectively. Bovine Serum Albumin (BSA) (as the standard in protein analy- Figure 6. Concentrations of volatile fatty acids (VFA) obtained in the effluents after biochemical methane potential (BMP) assays (first load and second load) for the diatom P. tricornutum and supercritical-fluid-extracted P. tricornutum.
Finally, it seems that metal content provided by the own diatom does not apparently affect the AD process. The use of microalgae (including diatoms) in co-digestion with rich carbon substrates is common for obtaining adequate nutrients and metal balance for the anaerobic system [68].

Biomass and Chemicals
The diatom Phaeodactylum tricornutum (CIB-41 from CIBNOR (https://www.cibnor. gob.mx/, accessed on 22 November 2021) La Paz, Mexico) was obtained from the Mexican Company "Microalgas Oleas de México S.A." (Guadalajara, Mexico). The diatom was cultured in F/2 culture medium as reported previously by Guillard [69] and harvested at the end of the exponential phase. The microalga (diatom) was provided freeze-dried (freeze-dry system Labconco Freezone 18 L Benchtop Freeze Dry System, Kansas, MO, USA), packed in vacuum-sealed plastic bags and then was stored at 4 ± 2 • C in the dark until use. All solutions used in this study were of analytical grade except for the standard solutions and chromatographic solvents. For supercritical fluid extraction, carbon dioxide and ethanol were purchased from Indura Air Products Group (Santiago, Chile) and Sigma-Aldrich (Santiago, Chile), respectively. Bovine Serum Albumin (BSA) (as the standard in protein analysis) and glucose (as the standard in carbohydrate determination) were acquired from Sigma-Aldrich (Santiago, Chile). For fucoxanthin identification and quantification, a standard of >99% purity (Sigma-Aldrich, Santiago, Chile) was used. Ethyl acetate, acetonitrile, and ultrapure (MilliQ) water were of chromatographic grade (Sigma-Aldrich, Santiago, Chile) for high-performance liquid chromatography (HPLC).

Determination of Particle Size Distribution of the P. tricornutum Freeze-Dried Biomass
The particle size distribution of lyophilized diatom powder was determined using an electromagnetic sieve shaker (CISA Sieving Technologies, BA200N model, Barcelona, Spain). Eight sieves with ISO 3310-1 (ASTM E11) sizes of 10,12,16,18,35,60,120, and 230 were used. After 15 min shaking, the material retained in each sieve and the bottom pan was weighed. The mean particle diameter (Dp) of the P. tricornutum powder was determined according to ASABE [70] using Equation (1).
where D P is the mean particle diameter (mm); d i is the diameter of the sieve opening i (mm); d i+1 is the diameter of the sieve opening above sieve i (mm); W i is the retained mass (g); and n is the total number of fractions. The samples were kept in sealed plastic bags in a refrigerator till further analysis.

SFE
The SFE experiments were performed using a commercial SFE unit (Spe-ed SFE Helix, Applied Separations, Allentown, PA, USA), which was described in detail by Salinas et al. [71]. The aim of this experiment was to evaluate the effect of extraction conditions such as the temperature of 30 • C and 50 • C; the pressure of 20, 30, and 40 MPa; and the concentration of ethanol as a co-solvent from 10% to 50% v/v on the extraction yield, fucoxanthin purity, and fucoxanthin recovery from the P. tricornutum biomass. Initially, different temperature and pressure conditions were evaluated using the freeze-dried biomass to optimize temperature and pressure parameters. For each extraction assay, 2.0 g of P. tricornutum biomass was packaged in a 24 mL extraction vessel (nominal volume), with the dimensions of 0.56 inch (1.4224 cm) of internal diameter and 5.9 inches (14.986 cm) of internal height (ASI Part Number 6414/7975; Applied Separations, Allentown, PA, USA), occupying a volume of 2.54 mL, resulting in a bed density of 0.787 g/mL, which is approximately 10.6% of the total volume of the extraction vessel. The empty space of the extraction vessel was filled with 1.0 mm glass beads. The extraction vessel containing the biomass was assembled into the heated jacket at a selected temperature and kept for 15 min until the desired temperature was stabilized. CO 2 was pumped until the desired pressure was reached, and the system was maintained for 10 min (static time) and the total time of extraction was set at 60 min according to a previous report on the extraction of carotenoids (xanthophyll) from microalgae [30]. The CO 2 flow rate was measured using an electronic flow meter installed at the exit of the sample collection flask. After the static time, the outlet valve of the bed was opened, and the micrometric valve was adjusted to reach a CO 2 flow rate of 2 L/min that corresponds to a mass flow rate of 3.62 g/min. At this mass flow rate, superficial CO 2 velocities (U: mm/s) through the extraction vessel were calculated according to the temperature (30 • C and 50 • C) and pressure (20, 30, and 40 MPa) of each treatment.
A factor that critically affects the cost of SFE processes for solid substrates is the extraction time [72], which in turn affects extraction yield that depends on substrate pretreatment, particle size (Dp), CO 2 temperature (T), pressure (P), and superficial velocity (U) [73]. This flow of CO 2 during 60 min corresponds to a ratio of solvent to feed (S/F) of 108.6. The effect of operative conditions on the yield and extraction of fucoxanthin was expressed in terms of the total yield of extract and fucoxanthin purity and recovery from P. tricornutum, respectively. These variables were calculated using the following Equations (2)-(4): Total Yield (%) = (We/Wb) × 100 (2) Purity (mg/g) = Wc/We Recovery (%) = (Wc/Wt) × 100 (4) where We is the weight of the extract (g), Wb is the weight of biomass (g); Wc is the weight of the fucoxanthin extracted (mg), and Wt is the reference weight of the fucoxanthin from conventional extraction (mg). This value was calculated on the basis of the initial content of fucoxanthin pigment in the diatom, expressed as mg of extract per gram of dry weight of P. tricornutum (equals to 9.82 ± 0.70 mg/g). Each experimental condition was replicated three to five times, and standard deviation (SD) was calculated for each value.

Fucoxanthin Quantification by HPLC
Fucoxanthin was extracted from 50 mg of supercritical fluids extracts, and methanol (0.05% w/v BHT) was added in all samples. The aliquots were filtered (Ø = 0.22 µm filter) and transferred into an amber vial for chromatographic analysis. Subsequently, fucoxanthin was separated and identified using the HPLC system, model 7100 (Merck Hitachi LaChrom, Tokyo, Japan) equipped with three pumps (flow rate of 1 mL/min), and a UV-Vis detector was used at 450 nm. The samples (20 µL) were injected into a capillary column (C18, 250 mm × 4.6 mm, 5 µm, Restek, Bellefonte, PA, USA). The mobile phase consisted of solvent A (ethyl acetate) and solvent B (acetonitrile:water 90:10). The gradient procedure was 100% solvent B for 10 min; 50% solvent B for 4 min; 40% solvent B for 2 min; and 100% solvent B for 1 min. Fucoxanthin was identified by comparing the retention times with that of the reference standard. Fresh standard solutions (1ppm to 20 ppm of fucoxanthin) were prepared and injected to generate the standard fucoxanthin curve. The calculation of the free fucoxanthin concentration in the samples was performed using the standard curve that was found to be linear over the required range (1-25 µg/mL; R 2 = 0.9971). All analyses were performed in triplicate.

Characterization of Biochemical Composition
The characterization of biochemical composition was performed from the original biomass, the whole P. tricornutum, and the exhausted biomass. The exhausted biomass was a unique sample composed of a mixture resulting from the biomass pool after all SFE extractions studied. The protein content of P. tricornutum was analyzed colorimetrically using a microplate reader (Synergy HTX Multi-Mode microplate reader, software Gen5 2.0, BioTek Instruments, Winooski, VT, USA) and BSA solutions for the standard curve (0.0−2.0 mg/mL) by following the Lowry method [74]. The results are expressed as the percentage of total proteins with respect to their dry biomass (% w/w). Total carbohydrate content was determined according to the modified protocols of Dubois et al. [75] and Geresh et al. [76] using a series of glucose solutions as the standard (0.0−1.0 mg/mL). The biomass used for carbohydrate measurement was subjected to acid hydrolysis for cell rupture. Briefly, 20 mg of the dry biomass was incubated in 5 mL of 2.5 M HCl for 3 h at 100 • C (Spectroquant Thermoreactor TR 320, Merck, Darmstadt, Germany). This mixture was neutralized with 5 mL of 2.5 M NaOH to obtain the microalgal extract. Finally, aliquots of 278 µL of extract, 167 µL of phenol solution (5% w/v), and 1000 µL of concentrated sulfuric acid were incubated for 30 min at room temperature, and the absorbance of these samples was measured at 483 nm using the microplate reader. The results are expressed as the percentage of total carbohydrates with respect to the dry biomass (% w/w). Total lipids were obtained according to the method of Axelsson et al. [77]. The dry biomass of P. tricornutum (~20 mg) was resuspended in chloroform:methanol (2:1, v/v) by manually shaking the tube vigorously for a few seconds or until the biomass was dispersed in the solvent system. Finally, 0.73% (w/v) NaCl water solution was added to produce a 2:1:0.8 system of chloroform:methanol:water (v/v/v). The previous evaporation with N 2 was performed using Flexivap Workstation (Glas-Col 109A YH-1, Terre Haute, IN, USA). Lipid content was expressed as the percentage of total lipids with respect to the biomass (% w/w). Proteins, total carbohydrates, and total lipids are expressed as the average of a triplicate (n = 3), and the data are expressed as mean ± SD.

Analytical Methods
The parameters pH, conductivity, total alkalinity (T-Alk), total solids, volatile solids (VS), mineral solids, total chemical oxygen demand (TCOD), soluble chemical oxygen demand, and volatile fatty acids (VFA) were determined according to the Standard Methods of the APHA et al. [78]. A Shimadzu model TOC-VCSH carbon analyzer was used for the determination of total organic carbon by the catalytic oxidation of total carbon present in the sample to CO 2 and detection by infrared spectrophotometry. Metal content was determined by flame photometry [78] with a Perkin-Elmer A Analyst 300 atomic absorption spectrophotometer. The methodology proposed by the US Department of Agriculture and the US Composting Council [79] was used for the determination of ammoniacal nitrogen (N-NH 4 + ), total Kjeldahl nitrogen (N-TKN), and phosphorus (P-P 2 O 5 ) content. Volatile fatty acids (VFA) such as acetic, propionic, butyric, isobutyric, valeric, and isovaleric acids were determined using a Hewlett-Packard HP-5890 gas chromatograph equipped with a 15 m × 5.3 × 10 −4 m (i.d.) Nukol-silica semi-capillary column and a flame ionization detector. Table 6 summarizes the main characteristics of the inoculum used for the biochemical methane potential (BMP) tests. The inoculum used was a baker's yeast vinasse obtained from La Golondrina plant, the municipal water company (EMACSA), Córdoba, Spain. The AD of P. tricornutum was assessed by performing BMP assays in batches. The 1.2 L reactors or digesters were maintained at mesophilic temperature (35 ± 2 • C) using a thermostatic water bath (LAUDA RTM 20) and were continuously agitated at 200 rpm. An inoculum-to-substrate ratio was maintained at 4 (on a VS basis). A total of 5.7 g inoculum of VS was added in each reactor using an effective volume of 1000 mL. The headspace of each flask was flushed with nitrogen to displace O 2 and promote AD conditions. The CH 4 produced was measured using a 1 L Boyle-Mariotte reservoir connected to each digester. NaOH solution (6N) was used assuming that the non-captured gas from biogas was methane. The volume of CH 4 was measured by water displacement. All methane volumes and yields were measured under standard temperature and pressure conditions (STP): 0 • C and 1 atm.

Anaerobic Inoculum and BMP
Before the experiments, the biomass was activated with GAL solution, a mixture of glucose (50 g/L), lactic acid (21 mL/L), and sodium acetate (25 g/L) with a TCOD concentration of 86,900 ± 75 mg O 2 /L and a pH of 7.05 ± 0.01. The experiments were performed in duplicate. Subsequently, the two digesters were fed with diatom P. tricornutum and the other two digesters with the extracted P. tricornutum (after the-SFE) to obtain CH 4 yields from both substrates.

Statistical Analysis
The Statgraphics Centurion XVIII ® Statistics software (StatPoint Technologies, Inc., Warrenton, VA, USA) was used for data elaboration and statistical analysis using a significance level of 95%. One-way analysis of variance, together with Tukey's test, was used for group extracts based on statistically significant differences. The effect of each factor and its statistical significance for each response variable were analyzed. A p-value of <0.05 was considered statistically significant. All experiments were performed in triplicate (n = 3), and the data are expressed as the mean ± SD.

Conclusions
In this study, we evaluated the use of P. tricornutum as biomass in the algae biorefinery concept using SFE, a green method, and biogas production to close the loop in a circular schedule. Fucoxanthin was the first valuable molecule to be recovered due to its many possible health benefits and high food market value. The best operating SFE parameters for the extraction yield, purity, and recovery of fucoxanthin from P. tricornutum by SFE were assessed. These optimal conditions were 30 MPa pressure at 30 • C and 30% v/v ethanol for extraction yield and 30 MPa pressure at 30 • C and 40% v/v of the extractant for fucoxanthin purity and recovery. Fucoxanthin recovery improved by 3.5 times at 30 MPa and 30 • C in the presence of ethanol, the most significant variable of SFE. Moreover, the diatom P. tricornutum was confirmed to be a potentially valuable natural resource for nutritional or nutraceutical purposes due to its good biochemical composition. The use of P. tricornutum and extracted diatom for AD enhanced the benefit of obtaining bioenergy in the form of biogas, obtaining methane yields of 77.15 ± 3.85 LSTP CH4/kg VS and 56.66 ± 1.90 LSTP CH4/kg VS for the whole diatom and the extracted P. tricornutum, respectively, in addition to fucoxanthin extraction; thus, implying its use a circular economy strategy, minimizing the potential costs associated with SFE as a costly pretreatment process for biogas production.
Based on our findings, P. tricornutum can be a useful candidate to diversify its multipurpose use from biotechnological and bioactive applications to sustainable energy production, playing a key role in the "algae biorefinery" concept.


text:
Production of Chicken Patties Supplemented with Cantaloupe By-Products: Impact on the Quality, Storage Stability, and Antioxidant Activity

This study investigated the effect of supplementation with cantaloupe peel (CP) and seeds (CS) (3, 6, 9, and 12%) powder on the quality and antioxidant activity of raw and cooked chicken patties during storage (-20°C/3 months). The addition of CP and CS powder increased protein, fat, ash, and fiber values of chicken patties compared with control, while carbohydrate, pH, and TBA were decreased at zero time and after 3 months of storage. The WHC, cooking yield, fat retention, and moisture retention were increased by increasing CP and CS powder addition ratios, while cooking loss and shrinkage were decreased. Also, CP and CS powder improved antioxidant activity, microbiological quality, and overall acceptability of chicken patties. The hardness of raw and cooked chicken patties was decreased with increasing CP and CS addition ratios. It is recommended to use CP and CS powder as functional ingredients in the preparation of functional foods.

Introduction
The by-products of fruits such as peels, seeds, and unused flesh are usually wasted and disposed of its. These agroindustrial residue wastes are a serious problem that must be managed effectively, and the fruits are natural sources of active compounds like vitamins, total phenolic, dietary fibers, and carotenoids [1,2].
In recent years, research has focused on peels and seeds of fruits due to their important nutritional and medicinal properties. In addition, the vegetable oil extracted from seeds has an excellent source of biologically active compounds and antioxidants [3,4].
Cantaloupe melon (Cucumis melo L.) is one of the most consumed crops all over the world because of its sweetness and delicious flavor; it belongs to Cucurbitaceae, a family which includes a large group of species with great economic importance. Often, cantaloupe peel and seeds that are agroindustrial residue waste products are eliminated and are a rich source of phytochemicals such as polyphenol compounds, carotenoids, and other substances that have a posi-tive effect on health. Also, these polyphenols have antioxidant activity as well as work to prevent the oxidation of fats and protect cells from damage by reactive oxygen species (ROS) [5,6].
Cantaloupe peel is one of the by-products that are eliminated. Several studies have indicated that the peel is rich in phenolic compounds, flavonoids, carotenoids, and other bioactive components that have a positive effect on health [7]. Furthermore, Goulas and Manganaris [8] reported that the peel contains had higher phenolic compounds and vitamin C contents than the pulp. Also, Mallek-Ayadi et al. [1] reported that the cantaloupe peel contained carbohydrates (69.77%), ash (3.67%), total dietary fibers (41.69%), and antioxidants like polyphenols and flavonoids (332 mg/100 g extract and 95.46 mg/100 g extract, respectively). In addition, it is considered as a good source of minerals, such as calcium, potassium, and magnesium.
Cucumis melo L. seeds are an excellent sources of protein, lipids, and fiber. Also, they are rich sources of minerals especially magnesium, phosphorus, sodium, and potassium. In Arab countries, the seeds are used after salting and roasting as a snack, as well as being dried and used as a flavoring agent for Indian dishes and sweets [9,10]. It is also a rich source of antioxidants and is also used to maintain shelf life [11]. In addition, Mansouri et al. [12] reported that the seed kernel of Cucumis melo is rich in unsaturated fatty acids like linoleic and linolenic as it contains 40-50% of the fatty acids and 20%-30% of the protein; it is considered as a good source of essential amino acids, such as isoleucine, methionine, tyrosine, phenylalanine, and valine [13][14][15]s, and this depends on the variety of Cucumis melo. As well, it were found that fatty acids ranged in oil from 41.6% to 44.5%. Also, it is a good source of fiber, minerals, and biologically active compounds [16,17]. It is a new food ingredient. Currently, it is used in some foods such as bakery products [18].
Nowadays, the meat industry is interested in replacing with synthetic antioxidants such as BHT, BHA, and TBHQ, etc which are toxic and harmful to health with natural products of plant origin that contain biologically active compounds with their health benefits [19,20]. And scientific evidence confirmed that the use of natural antioxidants would reduce fat oxidation [21], especially in chicken products most prone to deterioration.
Poultry meat is one of the most consumed meats in the world due to its availability and cheap price. It is an excellent source of biological protein, minerals, and vitamins and has low fat content, in addition to being rich in unsaturated fatty acids that can lead to oxidation, which leads to a decrease in meat quality and therefore low acceptance to its consumer [22].
Lipid oxidation and color are the most important factors that attract consumers to accept meat and its products; several studies have noted a decrease lipid oxidation in different meats, such as pork, beef, lamb, chicken, and goat after adding oregano, sage, rosemary, thyme, marjoram, caraway, basil, ginger, kinbow, pomegranate, cereal, walnut, and seaweed [19,[23][24][25][26]. It is worth to mention that a diet containing meat is rich in energy, saturated fatty acids, and cholesterol and poor in dietary fiber. The diet must contain fiber and incorporate it into food products to take advantage of important functional properties and technology [27][28][29]. Therefore, the aim of this study is to investigate the effect of supplementation of chicken patties by cantaloupe peel and seeds powder on the quality, storage stability, functional properties, antioxidant activity, and overall acceptability.

Materials and Methods
2.1. Materials. Fresh cantaloupe (Cucumis melo L.) fruit and fresh chicken breast meat, eggs, bread crumbs, salt, spices mixture (ground black pepper, ground cumin, and onion powder), and corn oil were obtained from a local market in Alexandria, Egypt. Grease-proof paper, polyethylene bags, and foam plates (22 × 17 cm) were purchased from Alexandria Local Market, Egypt.

Technological Methods
(1) Preparation of Cantaloupe Peel and Seeds Powder. Cantaloupe fruit was washed thoroughly with tap water; then, peel and seeds were separated from the pulp fruit using a knife. The peels were shredded into small pieces. Peels and seeds were dried in a hot air oven dryer at 45°C for approximately 16-18 hours till its moisture content reached 8%, then ground into a fine powder with an electric mill (Moulinex, MC3001), then packed into polyethylene bags, and kept at -20°C until used.
(2) Preparation of Chicken Patties. Chicken patties were prepared according to the method described by Nardoia et al. [30] with some modifications, by mixing of minced chicken meat, whole egg, bread crumbs, spices mixture, and salt ( Table 1). The bread crumbs in the formulas were replaced with 3, 6, 9, and 12% of cantaloupe peel (CP) or seeds (CS) powder. The chicken patties formulas were homogenized in a Braun Cutter Machine (Combi Max 700, USA) and then formed and processed into chicken patties (50 g weight, 10 cm diameter, and 1 cm thickness). A plastic packaging film was used to help maintain the shape of the chicken patties and kept at -20°C for 3 months. The samples were analyzed on zero time and after 3 months of frozen storage.

Cooking
Procedure of Chicken Patties. Chicken patties samples were grilled in a non-sticky pan (electric pan) with no added fat for 5 min on one side and 3 min for the other side, then cooled to room temperature (22 ± 3°C) as described by Mohamed [43]. One and a half milliliters of Folin-Ciocalteu's reagent (diluted 10 times) and 1.2 mL of Na 2 CO 3 (7.5% w/v) were added to 300 μL of water-soluble extract. Mixtures were shaken and kept at room temperature for 30 min before measuring absorbance at 765 nm using a spectrophotometer (Pg T80+, England). TPC was expressed as gallic acid equivalent in milligrams per milliliter of extract.  [45]. One milliliter of extract was mixed with 2.5 mL of phosphate buffer (0.2 M, pH 6.6) and 2.5 mL of potassium ferricyanide (1% w/v), and then, the mixture was incubated at 50°C for 30 min. After incubation, 2.5 mL of TCA (10% w/v) was added and the mixture was centrifuged at 1650 rpm/10 min. Finally, 2.5 mL of the supernatant solution was mixed with 2.5 mL of distilled water and 0.5 mL of FeCl 3 (0.1% w/v) and the absorbance was measured at 700 nm using a spectrophotometer (Pharmacia, USA). The FRAP values, expressed in milligrams of ascorbic acid per 100 mL, were derived from a standard curve.
2.9. Microbiological Evaluation. Total plate count (TPC) was determined for the samples at zero time and after 3 months of frozen storage (-20°C) by using pour plate method and plate count agar as medium according to ISO 8443 [46]. For coliform group bacteria, pour plate procedure and Violet Red Bile Agar medium were used according to ISO 4832 [47]. Regarding yeasts and molds, they were determined by plating 0.05 mL of diluted sample on potato dextrose agar (Oxoid CM) and incubated for 5 days at 25°C; yeast and mold colonies were counted separately according to ICMSF [48].
2.11. Texture Profile Analysis. Texture profile analysis of different chicken patties samples were determined by a universal testing machine (Cometech, B type, Taiwan) provided with software. An aluminum 25 mm diameter cylindrical probe was used in a "Texture Profile Analysis" CT V1.2 Build 9 (TPA) double compression test to penetrate to 50% depth, at 2 mm/s speed test. Hardness (g/s), springiness (mm), cohesiveness (ratio), gumminess (g/s), chewiness (mJ), and resilience were calculated from the TPA graphic.
2.12. Sensory Evaluation. Color, taste, odor, texture, and overall acceptability of cooked chicken patties were evaluated using 10 trained panelists from Food Technology Research Institute Agricultural Research Center, Alexandria. A 9-point hedonic scale was used (9 = like extremely and 1 = dislike extremely) according to Meilgaard [50].
2.13. Statistical Analysis. The statistical analysis was performed using one-way analysis of variance (ANOVA) using SAS statistical analysis software [51]. Means were compared by Duncan's test at the significance level of p < 0:05.  [52] found that the cantaloupe seeds flour contains 2.64% moisture, 17.64% crude protein, 4.12% ash, 35.48% total dietary fiber, and 30.43% fat. Also, results revealed that CP had the highest content in some minerals such as calcium (1000 mg/100 g) and potassium (940 mg/ 100 g). On the other side, the CS contains high amounts of magnesium, iron, zinc, and sodium (900, 2.50, 2.10, and 289 mg/100 g), respectively. The data obtained in the present study are similar mostly with those reported by Mallek-Ayadi et al. [1] and Mallek-Ayadi et al. [53]. Data in Table 2 showed that the higher water retention capacity (5.96 g/H 2 O) was observed with CP compared to CS (4.43 g/H 2 O). This might be due to the higher fiber content of CP. Results cleared that the oil retention capacity of peel powder is similar to that of seeds powder. These results are in line with those reported by da Cunha et al. [52]. Currently, the property of fat retention capacity was exploited in foods, especially meat products, which are lost during the cooking process, and this is likely to be useful to improve flavor and yield [54].

Antioxidant Activity of Cantaloupe Peel and Seeds
Powder. In terms of the antioxidant activity of CP and CS (Table 2), results revealed that CP was significantly (p < 0:05) higher in total phenolic (1050 GAE mg/100 g), DPPH (91.34%), and FRAP value (700 mg AAE/100 g). These results are less than those reported by Vella et al. [55] who reported that the total polyphenol content of cantaloupe peel and seeds were 25.48 and 1.50 mg GAE/g while FRAP values were 12.27 and 0.31 mg AAE/g, respectively. This may be due to the difference in the variety, degree of maturity, and environmental factors like geographic climate.

Proximate Composition of Raw and Cooked Chicken
Patties and Frozen Storage. The changes of chemical composition in raw and cooked chicken patties at zero time and after 3 months of storage at -20°C are given in Tables 3  and 4. At zero time, the moisture contents of raw and cooked chicken patties were decreased with higher addition ratio of CP and CS powder compared to control. At the end of storage period (3 months), the lowest moisture contents of raw chicken patties were observed with the samples containing 12% of CP or CS. These results are in agreement with Sharma and Yadav [56] who reported that the addition of pomegranate peel and aril bagasse powder caused a significant decrease in moisture content of chicken patties. Also, Mahdavi et al. [57] found that the moisture content of chicken burger was decreased in all chicken burger samples with increasing frozen storage time.
Also, the data in Tables 3 and 4 stated that the protein content was significantly higher in raw samples containing CS (3, 6, 9, and 12%) compared with the control sample at zero time, while higher protein content was observed with the sample containing 12% CP. Meanwhile, after 3 months of storage, the protein contents were significantly (p < 0:05) increased in all cooked samples compared with the same samples at zero time. These results are expected, since cantaloupe seeds powder had higher protein content (27.53%) than cantaloupe peel (7.50%) as indicated in Table 2. On opposite to our results, Sharma and Yadav [56] found that there were no significant (p > 0:05) differences in protein content of chicken meat incorporated with pomegranate peel and aril bagasse powder. Fat content in raw chicken patties was significantly increased with the increase of CS addition. The raw and cooked samples containing CS (12%) showed higher fat contents compared to raw and cooked control samples (Tables 3 and 4). Generally, frozen storage did not affect the fat content except in the sample containing 12% CS. This may be the result of cantaloupe seeds powder which are rich in lipids [52].
Furthermore, fiber content was significantly (p < 0:05) increased by increasing the addition ratio of CP and CS in raw and cooked samples at zero time and after frozen storage (3 months at -20°C). These results are in agreement with Mallek-Ayadi et al. [1] and da Cunha et al. [52], who reported that the cantaloupe peel and seeds flour were a good source of dietary fiber.
At zero time and after storage period (3 months), the ash content in raw and cooked chicken patties was significantly (p < 0:05) increased with the increasing ratio of CP and CS addition in all samples except the sample containing 3% CP compared with control (Tables 3 and 4). This could be due to the higher amount of ash of cantaloupe peel and seeds powder ( Table 2). The ash contents of cooked control samples and some CS cooked samples (CS 3%, CS 6%, and CS 12%) were significantly increased during storage, while all CP samples (raw and cooked) were not changed during storage. These results are in agreement with Sharma and Yadav [56].
Concerning the carbohydrate contents in raw and cooked chicken patties samples (Tables 3 and 4), the results showed a significant (p < 0:05) decrease in carbohydrate (%) with the increasing ratio of CP and CS whether at zero time or after 3 months of storage at -20°C. On the whole, frozen All determinations were carried out in triplicate and mean value ± SD. Different letters in rows indicate significant different value at p < 0:05.  International Journal of Food Science  Regarding to energy values in Figure 1, it was observed a significant (p < 0:05) increase in total calories (energy value) with the increasing in CP and CS addition in the cooked chicken patties either at zero time or after 3 months of frozen storage. Frozen storage had a significant (p < 0:05) effect on the energy values especially in cooked samples containing 3 and 9% CP and samples containing 3, 9, and 12% CS, whereas the energy values of all treatments were significantly (p < 0:05) increased by the end of the frozen storage period. These results were due to the decrease in moisture contents in frozen samples which resulted in an increase in protein and fat contents. Figure 2 indicated that at zero time, the pH values of raw patties (samples containing 9 and 12% CP and CS) and cooked patties (samples containing 12% CP and 9 and 12% CS) were significantly (p < 0:05) decreased compared to those of the control sample. Likewise, there was a significant decrease in all samples of raw patties and cooked patties samples containing 3, 6, 9, and 12% CP and 9 and 12% CS after 3 months of storage. Furthermore, the pH values had a significant (p < 0:05) increase after 3 months of frozen storage. The decrease in pH values with increasing ratio of the CP is due to the acidic nature of cantaloupe peel. Also, the increase in pH during frozen storage is due to the breakdown of protein, mainly amines. Similar findings were obtained by Chappalwar et al. [58]. They found that the addition of lemon peel powder to the chicken patties led to a decrease in the pH values, which may be attributed to the presence of polyphenols and flavonoids in lemon albedo as hesperidin, eriocitrin, and naringin. Figure 3 showed that at the zero time, the lower TBA values were found with raw patties samples containing 12% CP and CS and cooked samples containing 6, 9, and 12% CP and 9 and 12% CS compared to the control sample. Also, after 3 months of storage at -20°C, the raw and cooked patties samples containing CP and CS had lower TBA values compared to the control sample. This may be due to the effect of polyphenols and flavonoid compounds in CP and CS powder as antioxidant agents. Generally, TBA values in all patties samples were significantly (p < 0:05) increased with frozen storage (-20°C). This may be due to lipid oxidation and the formation of volatile basic nitrogen [59]. TBA values were within the permissible limits according to [60].

Thiobarbituric Acid (TBA). The results in
These findings are in agreement with Malav et al. [59], who found that the TBA values in all patties samples containing cabbage powder were significantly (p < 0:05) decreased compared to those in the control sample, which may be due to the presence of phenolic compounds in cabbage.
Also, Baioumy and Abedelmaksoud [61] found that formulating 5% orange peels (albedo) in the beef burgers has a positive impact; TBA values of the control and treatments were affected by the use of orange peel as there was a decrease in level of lipid oxidation, compared with the control sample. This confirms the positive effect of the orange peel on the quality characteristics and shelf life of beef burgers and reducing the microbial load during frozen storage.   International Journal of Food Science are due to protein denaturation and the release of water and fat from the beef burger [62]. Table 5 shows the changes in water holding capacity (WHC), cooking loss, cooking yield, thickness, and shrinkage in different chicken patties treatments at zero time and after 3 months of storage at -20°C. WHC significantly (p < 0:05) increased by increasing the ratio of CP and CS addition as well as frozen storage in all chicken patties treatments. WHC was increased from 1.17 cm 2 /0.3 g in the control sample to 1.62 cm 2 in the sample containing 12% CP at zero time and from 2.33 cm 2 /0.3 g to 2.79 cm 2 /0.3 g after frozen storage for 3 months for the same samples. This effect could be attributed to the presence of dietary fiber in CP. These results are in agreement with those reported by Serdaroğlu et al. [41] and Sharma and Yadav [56].
The increase in WHC values of chicken patties by frozen storage might be attributed to protein denaturation and loss of protein solubility [63].
Cooking loss was significantly (p < 0:05) decreased by increasing the ratio of cantaloupe peel and seeds powder addition; for example, the cooking loss was decreased from 18.07% with the control sample to 10.03% with the sample containing 12% CP at zero time and from 21.00% with the control sample to 12.19% with the sample containing 12% CP after frozen storage for 3 months. These results might be due to the ability of CP and CS to bind water and fat, which consequently decreased cooking loss. Haque et al. [64] found that the cooking loss of beef burger decreases with the addition of orange peel extract, and the cooking loss was increased at the beginning of storage and then decreased by the end of the storage period (after 60 days).
Data in Table 5 illustrated the cooking yield of chicken patties samples supplemented with CP and CS powder. The results indicated that the cooking yield was increased in all chicken patties samples containing CP and CS compared to the control sample at zero time and after frozen storage for 3 months. The decrement of cooking yield with  9 International Journal of Food Science frozen storage might be due to protein denaturation and loss of protein solubility which decreases water holding capacity consequently increasing moisture loss during cooking [63].
The change of diameter (shrinkage) was significantly (p < 0:05) decreased with an increasing ratio of cantaloupe peel and seeds powder. The higher shrinkage value was observed with the control samples at zero time and after frozen storage for 3 months (21.97% and 25.00%, respectively). Also, frozen storage had a significant effect on the shrinkage values which might be attributed to the ability of CP and CS to bind water and fat. Similar results were obtained by Bessar [65] who reported that increases in addition levels of orange and apple peels led to decreased shrinkage value in beef burgers.
No significant differences were observed in moisture retention among all treatments except samples containing 9 and 12% CP at zero time. Storage at -20°C for 3 months caused an increase in moisture retention for samples containing 3, 6, 9, and 12% of CP and 9 and 12% of CS. Also, findings indicated that fat retention was increased (p < 0:05 ) in sample patties containing 12% of CP and 3, 6, 9, and 12% of CS at zero time and after 3 months of storage at -20°C.
Meanwhile, reduction thickness was decreased (p < 0:05) by increasing the ratio of CP and CS in patty samples compared to the control samples at zero time and after 3 months of storage. Generally, frozen storage had an effect on moisture retention and shrinkage and thickness of patties, while it did not affect fat retention. These results were in line with Chappalwar et al. [58] who found that moisture and fat retention were increased with increasing levels of lemon albedo in chicken patties is due to the presence of fiber in lemon albedo, which has the ability to bind water and oil.
Hartmann et al. [66] who observed that the cooking yield and moisture retention were increased significantly in hamburger samples containing pumpkin peel flour (PPF). On the other hand, there was a decrease in the shrinkage percentage with the samples containing 3 and 4% PPF. This may be due to the presence of fiber in PPF which has the ability to interact with meat proteins by creating a network that prevents the transfer of water from the product to the surface, and this leads to an increase in cooking efficiency and reduces the shrinkage of the burger.

Antioxidant
Activity of Chicken Patties. The antioxidant activity of raw and cooked patties during storage at -20°C for 3 months are shown in Figure 4. The addition of CP and CS to cooked patties formula increased the total phenolic contents (TP). The TP contents were significantly (p < 0:05) increased with the increase of CP and CS addition ratio. The addition of CP caused increased total phenolic contents compared the CS, and the best TP content was observed with the sample containing 12% CP. The findings revealed that at zero time, the DPPH radical scavenging activity (%) of raw and cooked patties significantly (p < 0:05) increased by increasing the ratio CP and CS powder addition and significantly (p < 0:05) decreased after 3 months of storage. Ferric reducing antioxidant power (FRAP) was significantly (p < 0:05) increased by increasing of CP addition ratio. The raw and cooked samples containing CP powder showed a higher FRAP value than the CS and control samples.
Total phenolic contents also showed a similar trend of antioxidant power (FRAP) that is increased significantly (p < 0:05) by increasing the CP ratio in patties samples for raw and cooked patties. It could be noted the total phenolic contents and FRAP were higher in cooked patties compared with raw patties; this may be due to the low moisture content after cooking and also the patties which retained the content of phenolic compounds after cooking which is an indicator that this product has health benefits for the consumer [30]. No significant differences were observed in FRAP values among all containing CS compared with the control sample at zero time, while the FRAP value decreased significantly (p < 0:05) after 3 months of storage in samples containing CS. Moreover, total phenolic contents decreased significantly (p < 0:05) with frozen storage especially in cooked sample patties containing 3, 6, and 9% CP and 9 and 12% CS.

10
International Journal of Food Science  13 International Journal of Food Science 3.8. Microbiological Quality. For microbial load, i.e., total bacterial count (TBC), coliform group bacteria, and yeast and mold count (log cfu/g) in raw and cooked patties treatments during storage at -20°C for 3 months, the findings indicated that at zero time, the TBC of patties samples was decreased with increasing the ratio of CP and CS addition compared with the control sample (Table 6), while, after 3 months of storage, the TBC was gradually increased. The increase in TBC after 3 months of storage may be due to an increase of amino acids and fatty acids resulting from the hydrolysis of proteins and fats during storage which is suitable for the growth of microorganisms. In general, the TBC after 3 months of storage is less than the permissible limit which is log10 7 cfu/g for cooked meat products [67]. The coliforms were not detected in all patties samples at zero time and after 3 months of storage. This may be due to the high temperature during cooking that led to the destruction of the coliform bacteria. Similar results were obtained by Malav et al. [59]. Also, yeasts and molds were not detected in all patties treatments at zero time. Moreover, it could be observed that yeast and mold counts were decreased by increasing the ratio of CP and CS addition compared with the control sample after 3 months of storage. This may be due to the CP and CS which are rich sources of phenols and flavonoids, which have an antimicrobial effect [1]. Tables 7 and 8 shows the color values (lightness, redness, and yellowness) of raw and cooked chicken patties treatments at zero time and after 3 months of frozen storage. It could be observed that there was a significant increase in lightness (L * ) with a steady increase in the percentage of CP or CS in raw and cooked chicken patties. The highest lightness value was observed in patties containing 12% CS in raw and cooked patties by 14 International Journal of Food Science the end of storage period. The increase in lightness could be attributed to the color of CP and CS. These results are in agreement with those reported by Chappalwar et al. [58] who observed a significant increase in lightness by increasing the ratio of lemon albedo powder in chicken patties. Regarding redness (a * ) value, data revealed that using CP in raw and cooked chicken patties resulted in a significant increase in a * value compared to control sample during frozen storage period. On the other hand, the samples containing CS (3, 6, 9, and 12%) showed a significant (p < 0:05) decrease in redness after frozen storage for 3 months. However, there was a significant (p < 0:05) increase in redness value for cooked samples after frozen storage.

Color of Raw and Cooked Chicken Patties Treatments as Affected by Addition Ratio of CP and CS Powder during Frozen Storage. Data presented in
As for yellowness (b * ) value, the raw and cooked samples containing CP and CS were higher in yellowness value compared to the control sample, except raw and cooked patties samples containing 3% CS at zero time which showed the lowest yellowness value compared to control and other treatments. These findings might be because CP and CS were a good source of carotenoid pigments and polyphenol Table 6: Changes in microbiological quality of raw and cooked chicken patties supplemented with cantaloupe peel (CP) and seeds (CS) powder during storage at -20 for 3 months.

Treatments
Chicken patties

16
International Journal of Food Science  [66] observed that hamburger containing 3% pumpkin peel flour had significantly increased lightness, redness, and yellowness which might be due to the presence of compounds in the peel like chlorophyll, carotenoids, and flavonoids, which are natural colorants in fruits and vegetables.
3.10. Texture Profile Analysis of Raw and Cooked Chicken Patties Treatments as Affected by Addition Ratio of CP and CS Powder during Frozen Storage. Table 9 shows the texture profile analysis (hardness, gumminess, chewiness, springiness cohesiveness, and resilience) of raw and cooked chicken patties during frozen storage for 3 months at -20°C. The hardness of raw chicken patties was significantly (p < 0:05) decreased by increasing the ratio of CP and CS addition, and the higher decrease was observed with samples containing 12% CP or 12% CS powder at zero times compared with the raw control sample. This may be due to the increase in dietary fiber in the CP and CS powder, as the increase in fiber gives a softer texture to the product. Also, the results showed a significant difference in hardness values among all treatments after 3 months of storage except samples containing 3% CP and 3 and 6% CS powder. Likewise, there was a significant (p < 0:05) decrease in hardness value among cooked samples containing 6, 9, and 12% CP and 9 and 12% CS after 3 months of storage. Generally, the hardness values were increased significantly (p < 0:05) with the increase in frozen storage time.
The gumminess of raw and cooked chicken patties samples significantly (p < 0:05) decreased with increasing the ratio of CP and CS powder addition through frozen storage (Table 9). In general, gumminess values were increased slightly with increasing the frozen storage time. Similar results were obtained by Chappalwar et al. [58] who observed a decrease in hardness, gumminess, cohesiveness, and springiness values of chicken patties by increasing the ratio of lemon albedo powder. They attributed this to the effect of lemon albedo on the protein system, the presence of water, and the binding of fats in the lemon peel which provides a smooth texture.
Chewiness value also showed a similar trend of resilience that is increased with increasing the CP and CS ratio in patties samples. Moreover, during frozen storage, chewiness values were increased significantly (p < 0:05) expect samples containing 6 and 12% of CP powder. However, there was no significant (p > 0:05) difference in resilience value of raw and cooked samples after 3 months of frozen storage. The storage times did not affect the resilience value of all treatments. No significant (p > 0:05) differences were observed in springiness value among the control and samples containing CP and CS powder during frozen storage. On the other hand, the springiness values of cooked samples containing CP powder were significantly (p < 0:05) decreased during frozen storage, while the higher springiness values by the end of storage were found with samples containing CS powder. Cohesiveness values of both raw and cooked patties were significantly (p < 0:05) lower with samples containing 3, 6, 9, and 12% of CP and 12% of CS at zero time. Moreover, there was a significant (p < 0:05) decrease in samples containing 9 and 12% CP and 12% CS of raw patties and 12% CS in cooked patties after 3 months of storage.
Cohesiveness values among samples containing 9 and 12% CP were significantly (p < 0:05) increased after 3 months of frozen storage. Sharma and Yadav [56] found that incorporation of pomegranate peel powder (PPP) in chicken patties led to significant increase in hardness and gumminess, while chewiness value was lower in PPP-treated patties.
3.11. Sensory Evaluation. Sensorial evaluation of chicken patties is depicted in Figure 5. There were no significant (p > 0:05) differences in the color score found between the control sample and chicken patties containing CP and CS powder. Taste score was significantly (p < 0:05) decreased in patties samples containing 9 and 12% CP and 6, 9, and 12% of CS. This may be due to the association of the taste with high phenolic compounds in the peel, which caused a

18
International Journal of Food Science slightly bitter and acidic taste to the patties [58]. Odor and overall acceptability scores of 3% CS were significantly (p < 0:05) higher than those of the control sample. Moreover, there was a significant difference (p < 0:05) in texture score between control and samples containing 6, 9, and 12% CP and CS powder. The frozen storage had a negative effect on most sensory characteristics of among treatments. It was observed that the sample containing 3% CS was significantly (p < 0:05) decreased in texture and overall acceptability scores after 3 months of frozen storage. Hartmann et al. [66] observed that there were no significant (p > 0:05) differences in appearance, texture, color, and purchase intention between the control sample and hamburger containing at 1, 2, 3, and 4% pumpkin peel flour (PPF), while the control sample and sample containing 1% PPF were more accepted than 4% PPF in regard to overall acceptance. This may be due to the presence of phenols in the peel of vegetables, like tannins, and this enhances the astringent taste.

Conclusion
This study was carried out to improve the quality, cooking properties, and antioxidant activity of chicken patties by the addition of cantaloupe (peel and seeds) powder which was considered as a good source of phytochemical components, crude fibers, protein, fat, and minerals. The use of cantaloupe (peel and seeds) powder improved the functionality, quality properties, and antioxidant activity of chicken patties. Also, the results showed that the addition of CP and CS caused a significant (p < 0:05) increase in cooking yield, fat retention, and moisture retention. In addition, chicken patty samples had a high microbiological quality compared to the control sample. Chicken patties fortified with CP and CS powder at a ratio of 3% showed the best overall acceptability compared with the control sample and other treatments. Generally, this study recommended the use of CP and CS at a ratio of 9% in the development of meat products' with good functional properties and acceptability.

Data Availability
The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.