Formulation and Evaluation of Phyllanthus Wightianus-Based Active Packaging Film for Meat Preservation

Formulation and Evaluation of Phyllanthus Wightianus-Based Active Packaging Film for Meat Preservation | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Formulation and Evaluation of Phyllanthus Wightianus-Based Active Packaging Film for Meat Preservation Kangkang Xie, Samreen Ahsan, Amna Tariq, Muhammad Adil Farooq, and 7 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-3913167/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract A biopolymer-based edible film was prepared using Phyllanthus wightianus to extend the shelf life of beef patties. For this purpose, the film was prepared by using polyvinyl alcohol (PVA, 5% W/V), carboxymethyl cellulose (CMC, 1% W/V), glycerol (0.1% V/V) as a plasticizer, flaxseed gel and P. wightianus extract @ 0, 1, 2 and 3% in various treatments. The film was analyzed at the interval of 0, 10, 20, and 30th days at 25°C for thickness, the film's opacity, degree of solubility and swelling, water vapor permeability, and Staphylococcus aureus and Escherichia coli antibacterial activity. After a month of storage study, the developed film was used to coat beef patties and analyzed for antioxidants, pH, peroxidation, and moisture. A total plate count test was performed for antimicrobial analysis, and beef patties were further evaluated for sensory evaluation parameters. The results showed that film has substantial antimicrobial potential suitable for longer storage and helpful in delaying the spoilage of beef patties by controlling lipid peroxidation and microbial growth of meat spoilage bacteria, especially S. aureus and E. coli. In conclusion, the films developed with 3% P. wightianus extract and flaxseed gel prolong the shelf life of beef patties throughout storage. Biological sciences/Microbiology Physical sciences/Materials science/Techniques and instrumentation/Characterization and analytical techniques Phyllanthus wightianus Flaxseed gel Active film Antimicrobial packaging film Beef patties Storage study Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 1. Introduction The increased consumer awareness of synthetic packaging for food products has shifted the attention of researchers toward developing advanced eco-friendly packaging. Many concepts have been introduced in packaging, and "Active Packaging" (AP) is one of those. Active packaging modifies its physical state to improve its safety, sensorial, or shelf-life characteristics while preserving the food's quality. Intelligent food packaging, also known as "active packaging," functions as an additional purpose above conventional packaging 1 . Active packaging enhances the product's shelf life by interacting with the consumables inside the packaging. It has been classified into systems that either absorb undesired substances or release active substances. It covers a wide range of technologies, some of which, like oxygen scavengers and moisture absorbers, have a well-defined role in the excellent consumer market, while others, such as antibacterial materials, are still in the early stages of commercialization 2 . Active packaging gently releases the added active ingredients into the food product while absorbing harmful elements like moisture, oxygen, and free radicals to preserve the quality attributes of food 3 . The area of AP is growing substantially due to continuous changes in contemporary consumer demand and industry trends. The primary AP systems include those that produce carbon dioxide and ethanol, absorb and regulate moisture, scavenge oxygen, and work with migrating and non-migrating antimicrobial (AM) systems 4 . Edible packaging films with antimicrobial properties have generated a lot of scientific and technical attention recently because they have the ability to improve the safety and quality of food. There are two terms, "film" and "coating" used interchangeably for packaging systems however, film is defined as a stand-alone wrapping material, whereas a coating is a thin layer that is created immediately on the surface of food and has a thickness of less than 0.3 mm 5 . Antimicrobial active packaging film can be developed using antimicrobial substances, including organic acids, bacteriocins 6 , and extracts 7 . The AM agents inhibit by changing the microflora's cell wall composition and showing “cidal” or “static”. These compounds ordinarily include natural antimicrobials, antioxidants, antimicrobial polymer, and biotechnology products 8 . Phyllanthus wightianus is monoicous, sub-shrub up to 1m in loose whorls, and piliferous. Leaves alternate, distiches, oval, green above. All parts of the plant ( P. wightianus ) have been utilized for a long time as part of ethno- medicine, as an antidiarrheal, against jaundice, and to treat dieresis. It demonstrates many biological characteristics, including “antimicrobial”, pesticide, analgesics, tissue repair, and antioxidant capabilities. Flaxseed (Linumusitatissimum L.) is an individual from the variety Linum in the family Linaceae. Flaxseed is also named as functioning or bioactive food because it contains different supplements, such as bioactive compounds and alpha-linolenic acid 9 . In recent years, flaxseed (linseed) ( Linum usitatissimum ) has become a popular functional food ingredient. Flaxseed gel is a type of hydrocolloid widely used in the food sector. Increased neutral polysaccharide content, such as xylose, has been demonstrated to improve flaxseed gel rheological qualities by improving shear thinning and gelling properties. Because of its favorable hydrophilicity and remissibility, this gel has many applications in meat products 10 . The beef has a shelf life of around 3–5 days at 4 ºC. The high quantity of moisture, oxygen, and enzyme ease the lipid oxidation, and the high protein content prevents microbial growth during shelf life 11 . The current research emphasizes on shelf life enhancement of beef patties by using active edible packaging film. This film was prepared by using the antimicrobial properties of Phyllanthus wightianus and gelling features of flax seed as a core ingredient for its functional properties and then to determine the physicochemical, antimicrobial, textural, and sensory properties of edible active packaging film and beef patties. 2. Materials and Methods 2.1 Materials All the substances, i.e ., Carboxy methylcellulose, polyvinyl alcohol, citric acid, glycerol, and Mueller Hinton agar, were purchased from Sigma Aldrich and Merck Company. Phyllanthus wightianus and flaxseed were purchased from the local market of the city of Rahim Yar Khan, Punjab, Pakistan. 2.2 Preparation of flaxseed gel The flaxseeds were cleaned manually by removing any foreign material, then washed and soaked with distilled water. A magnetic stirrer was used to stir the flaxseed for 5 hours. Then the water mixture was placed in a water bath at 60°C at 300 rpm, filtered through muslin cloth, and assembled with minimal stirring using a glass rod 12 . 2.3 Preparation of Phyllanthus wightianus extract The dried leaves of Phyllanthus wightianus were used in a Soxhlet apparatus; the leave powder of Phyllanthus wightianus was taken in a thimble, and solvent hexane was used for extract extraction for 4 hours that may contain e.g. , Caffeic acid, catechol, and pyrogallol. The extract was filtered with Whatman filter paper to get a refined extract and placed in a rotary evaporator (Daihan scientific) at 35 ºC during the working evaporator bath temperature was 53ºC, and the chiller temperature was 4ºC for 1 hour 13 . The details are given in Fig. 1 . 2.4 Active packaging film preparation The film for active packaging was prepared by mixing Polyvinyl alcohol (5%) and carboxymethylcellulose (1%) in hot distilled water at 60ºC by constant stirring in the water bath (WB-22). Then glycerol (0.5%) was added as a plasticizer in a polyvinyl alcohol and carboxy methylcellulose solution with continuous stirring. After that, citric acid (5%) and flaxseed gel were added by following extract of P. wightianus (1%) was added in T 1 , (2%) in T 2, and (3%) was added in T 3 solution. A homogenizer was used to homogenize the solutions independently (HG-15D) for 5 minutes at 700 rpm. The prepared solution was transferred into each petri plate (15 mL) and placed in an incubator (model No. 30-1060) at 50ºC and 50% relative humidity for 24 hours. After drying, the intact films were carefully removed as a peel with the help of a spatula 14 . The details are provided in Fig. 2 . 2.5 Characterization of antimicrobial active packaging film 2.5.1 Thickness The film thickness was measured with a handheld micrometer cut into rectangular strips of 1× 5 cm 15 . 2.5.2 Film opacity The film opacity was calculated using a colorimeter calibrated by using white and black plates. The film's absorbance was evaluated using a UV spectrophotometer at a wavelength of 600nm. \(\text{O}\text{P} \left(\text{%}\right) = \frac{OPb}{opw } \times 1\) 00 O pb and O pw are opacity on at black and white background, respectively 16 . 2.5.3 Degree of swelling The film was cut into 2×2 pieces to assess swelling, solubility, and degree. The pieces dried at 105 ºC to a constant weight (M 1 ) to achieve initial dry mass. After weighing, the film (M 1 ) was put into 50 ml beakers with 30 ml distilled water and held at 25ºC for 24 hours. The wet films were dried with filter papers before being weight (M). The swelling degree is calculated by using this equation $$\varvec{F}\varvec{i}\varvec{l}\varvec{m} \varvec{s}\varvec{w}\varvec{e}\varvec{l}\varvec{l}\varvec{i}\varvec{n}\varvec{g} \varvec{D}\varvec{e}\varvec{g}\varvec{r}\varvec{e}\varvec{e}=\frac{\varvec{M}2-\varvec{M}1}{\varvec{M}1}\varvec{x} 100$$ Where M1 = Initial dry mass M2 = Final dry mass 17 . 2.5.4 Water vapor permeability Every film sample (4cm ×4cm) used to seal test tubes containing 10g of silica (0 percent RH). All test tubes were measure and immerse in a desiccator containing distilled water. The test tubes were tested again after 24 hours to see how much they had grown in weigh 14 . 2.5.5. Antimicrobial activity Disc zone of inhibition assays were carried out as a qualitative test for antibacterial activity. The films were cut into 10 mm discs aseptically and then placed on Muller Hinton plates for E. coli O157:H7 and Staphylococcus aureus , which had previously been inoculated with 0.1 mL of 10 5 CFUs/mL inoculum. Samples were incubated for 48 h at 37°C. The thickness (millimeters) of the inhibition zone around the film disk (colony-free perimeter) was measured with a millimeter scale, and the growth below the film disks (the contact area of edible film with agar surface) was visually examined 18 . 2.5.6. Antioxidant activity of antimicrobial active packaging film A different volume of film sample was individually reacted at room temperature for 1 hour in the dark with 4 mL of 100mol/L DPPH methanol solution. At 517nm, the absorbance of the reaction solution was measured. The absorbance of the reaction solution and the blank are A 0 and A 1 19 . DPPH radical scavenging activity (%) = \(A0-\frac{A1}{A0}\times 100\) 2.6 Analysis of Beef Patties coated with antimicrobial active packaging film 2.6.1 pH measurement A hand-held pH meter was used to calculate the pH of 10% of the homogenate sample after microbiological analysis 14 . 2.6.2 Peroxidation of Lipid Lipid samples were dissolved in a 30 mL mixture and glacial acetic acid (3:2, v/v). A saturated KI (1 mL) solution was added. The mixture was hand-shaken for 30 seconds before being held in the dark for another 5 minutes. The mixture was titrated against sodium thiosulphate (0.1 mol/L) after adding 100 mL distilled water until the yellow color almost vanished. After that, 1 mL of starch indicator solution (0.05%) was added. The titration was continued until the blue color completely vanished 20 . 2.6.3 Moisture content Each square film sample of 2cm×2cm was weighed for initial weight and dried at 105ºC in a drying oven for 24hrs 21 . 2.6.4 Total Plate Count (TPC) / Total viable count (TVC) The beef patties (10g meat) were removed aseptically and homogenized in a stomacher in sterile saline. Serial dilutions were performed, with sufficient dilutions on plate count agar (PCA) for total viable count (TVC). Counts were done at 5 days interval of chilled storage and represented as log cfu/g 14 , 22 . 2.6.5 Sensory evaluation Overall acceptability on the 9-point hedonic scale of beef patties was carried out to assess the acceptability of coating by a panel of 6 judges. The panels of judges were trained and familiar with products attribute. Scales of hedonic, where 9 is equal to extremely liked and 1 is equal to extremely disliked, were used. Water was provided for rinsing mouth after each sample 23 , 24 . 2.6.7 Statistical evaluation The information collected for every variable was subjected to statistical evaluation in order to measure significance level 25 . Two-factor factorial design under CRD was performed to determine results for all parameters by using statistics 8.1 software. 3. Results and Discussion Current research aimed to prepare antimicrobial edible active packaging film for beef patties by using Phyllanthus wightianus as an antimicrobial agent and flaxseed gel as a lubricating and functional ingredient. 3.1 Characterization of anti-microbial active packaging film 3.1.1 Thickness of anti-microbial active packaging film The thickness affects the mechanical properties, water vapour permeability, barrier characteristics, and transparency of edible films. Thinner films have better optical properties but decreased mechanical properties 26 . The thickness of the film plays a vital role in the suitability of utilizing the film for food packaging because it affects both the obstacles and mechanical features of the film. The thickness of a film is determined by its composition (solid content) and its processing parameters 14 . The results regarding the thickness of edible film depicted a non-substantial ( P > 0.05) impact of treatments and the effect of treatments and periods on each other. The overall effect of storage days showed a significant variation between (0.046 to 0.049 mm) during 30 days of storage (Table 2 ). Table 1 Effect of Treatment on active packaging film Treatments thickness (mm) Opacity (%) Swelling (%) Water vapour permeability (gm − 1 s − 1 pa 4 ) Zone of Inhibition (mm) of Staphylococcus aureus Zone of Inhibition (mm) of E.coli T o 0.0473 AB 26.00 C 5.45 B 1.92 B 0 0 T 1 0.0468 B 26.51 C 5.52 B 1.83 C 19.03 C 16.46 C T 2 0.0490 A 27.97 B 5.51 B 1.99 B 23.40 B 19.30 B T 3 0.0485 AB 30.32 A 5.83 A 2.08 A 25.56 A 22.1 A A−c Means in column with different superscripts differ substantially T 0 = CMC + PVA + CA + Glycerol + flaxseed gel T 1 = 1% P. wightianus + CMC + PVA + CA + Glycerol + flaxseed gel T 2 = 2% P. wightianus + CMC + PVA + CA + Glycerol + flaxseed gel T 3 = 3% P. wightianus + CMC + PVA + CA + Glycerol + flaxseed gel Table 2 Effect of storage on active packaging film Storage Period Days Parameters 0 10 20 30 Thickness (mm) 0.0495 A 0.0485 AB 0.0473 BC 0.0463 C Opacity (%) 27.7 A 27.75 A 27.76 A 27.52 A Swelling (%) 6.25 A 5.73 B 5.30 C 5.03 D water vapour permeability (gm − 1 s − 1 pa 4 ) 0.27 D 1.90 C 2.66 B 2.99 A A−D Means in column with different superscripts differ substantially The results of our finding were in correspondence with previous study 27 , which studied the influence of extreme alkaline pH on chicken protein edible film formation and found that lower film thickness was 68.8 µm, because of low moisture content. As a result, the control film had a thickness of 185.2 µm, which was much higher than the other groups. So, it can be stated that the loss of thickness in the current study during storage could also be due to moisture loss. The results of our findings were also similar to Mehdizadeh et al. 28 who have studied to increase the shelf life of beef by a chitosan-starch film incorporating pomegranate peel extract and Thymus kotschyanus essential oil. They have found that film thickness in chitosan-starch varied in the range from 0.046 to 0.063 mm. After that Utami et al. 29 studied the effects of cinnamon bark essential oil ( Cinnamomum burmannii ) on edible film characteristics and fresh beef quality. They have found that the thickness of edible films ranged from 0.114 to 0.176 mm, slightly higher than current research findings. 3.1.2. Opacity of anti-microbial active packaging film For light-sensitive food products and customer acceptability, the transparency and color of the food packaging materials are significant parameter 16 . The results regarding the opacity of edible film showed a non-significant ( P > 0.05 ) effect of days and the effect of treatments and storage on each other. However, the effect of treatment showed a highly significant ( p < 0.01 ) effect on the film, as given in Table 1 . The mean values for the opacity of the film are given in Table 1 . The results show that a minimum opacity value of 26.00% was recorded in T 0 . The overall effect of storage days showed non-significant variation between (26.00 to 30.32%) during 30 days of storage (Table 2 ). The resultant values of recent findings' opacity were higher than the previous reported study 14 which studied the development of water-resistant active packaging film prepared by carboxymethyl cellulose, polyvinyl alcohol, and aloe vera. They have found that film opacity ranges from 1.35 to 2.96% unit. Moreover, Kadam et al. 30 studied the influence of Nigella sativa seedcake polyphenolic extracts on the physicochemical qualities of chitosan-based films. They found that film opacity ranged from 3.65 to 16.75mm, which was lower than current findings. Likewise, Sun et al. 31 has studied the preparation and characterization of antimicrobial active food packaging film based on chitosan and incorporating apple peel polyphenols. They have found that film opacity ranges from 0.71 to 4.25 (A·mm − 1 ). Similarly, Rambabu et al. 21 has studied the mango leaf extract incorporated chitosan antioxidant film for active food packaging. They have found that film opacity ranges from 0.72 to 1.28 mm. For films containing various concentrations of CA, it was observed that the opacity increased with an increase in the concentration. Film opacity is an important parameter of the packaging film, as consumers prefer to be able to see the packed food items. 3.1.3. Degree of swelling of anti-microbial active packaging film The swelling capability of a film to hold water molecules defines its swelling capability. It has been reported that an increase in the concentration of citric acid usually causes decrease in the degree of swelling of films. Moreover, the swelling properties of film are inversely proportional to the degree of crosslinking, and if the crosslinking increases, then degree of swelling automatically decreases 14 . The results regarding degree of swelling of edible film showed highly substantial ( P < 0.01) impact on storage days and significant ( P < 0.05 ) effect of treatment variation however, non-significant effect of different concentrations and different storage days on each other (Table 1 ). The results show that after control sample the minimum value of degree of swelling 5.51% recorded in T 2 that was prepared with 2% P. wightianus while the maximum mean value was 5.83% in T 3 with 3% P. wightianus extract. The overall effect of treatment showed increase in between 5.45 to 5.83% from T 0 to T 3 and 6.25 to 5.04% decreases during 0 to 30 days of storage. The degree of swelling in a recent study was higher than that (Wulandari and Yuliatmo) 32 they studied the coating effect on edible film made from a bovine split hide gelatin on the characteristics of beef meatballs. They have found that coating concentration in soluble protein ranges from 0.178 to 0.346%. However, it was lower than the previous study 33 , which studied the physical chitosan properties of films containing naturally occurring antioxidants. They have found that the swelling degree of the chitosan-based film ranges from 191 to 233%. Likewise, Riaz et al. 34 have also studied the antimicrobial preparation and characterization of active food packaging film based on chitosan and incorporating apple peel polyphenols. They have found that swelling degree of chitosan-based apple polyphenol film ranges from 35.79 to 56.25% treatment containing chitosan- based polyphenol significantly increased the swelling degree. 3.1.4. Water vapour permeability of anti-microbial active packaging: Separating foods from ambient atmosphere vapour is one of the key functions of preservative films, which helps to avoid or delay food degradation. The water vapour permeability of the film should be kept as low as possible because it is primarily used to inhibit moisture from transferring from the food to the environment, hence extending the shelf life of food 34 . Water vapour permeability of films can be affected by a variety of parameters, including the chemical nature of macromolecules, crystallinity, molecular mass, orientation, and degree of cross-linking 35 . The results regarding water vapour permeability of edible film showed highly substantial ( p < 0.01 ) effect of storage period, treatments and for the effect of treatments and storage on each other. The effect of treatment showed substantial variation in between 1.83 gm − 1 s − 1 pa 4 (T 0 ) to 2.08 gm − 1 s − 1 pa 4 (T 3 ) and among storage days, it showed variation in between 0.27 to 2.995 gm − 1 s − 1 pa 4 during storage. The results of our findings were similar to previous study 36 who has studied on gelatin-coated paper for beef packing. They have found that water vapour permeability ranges from 0.2855 to 0.2635(g mm m- 2 h − 1 kPa − 1 ). However, the results of our findings were lower than Riaz et al. 34 they had studied the antimicrobial preparation and characterization antimicrobial active food packaging film based on chitosan and incorporating apple peel polyphenols. They have found that water vapour permeability of chitosan-based apple polyphenol film ranges from 14.65 to10.01 gm − 1 s − 1 pa 4 and treatment containing chitosan- based polyphenol significantly increased the water vapour permeability. Moreover, Kadam et al. 30 has studied the influence of polyphenolic extracts of Nigella sativa seedcake on the physicochemical properties of chitosan-based film. They have found that water vapor permeability o chitosan-based film ranges from 0.207 to 0.203 gm − 1 s − 1 pa 4 . 3.1.5 Antimicrobial Activity of anti-microbial active packaging film: Microbial contamination, which causes the spread of foodborne infections, is one reason that causes food modifications. As a result, regulating or preventing food contamination is crucial, and various ways have developed to achieve this goal. One of these techniques is to utilize antimicrobial active packaging, which interacts with the packaged food to extend its shelf life 37 . The results show maximum inhibition zone with T 3 treatment with 3% concentration of antimicrobial extract for S taphylococcus aureus and E. coli (Table 1 ). The Fig. 3 A and Fig. 3 B is also depicting zone of inhibition. Natarajan et al. 13 has studied the P. wightianus Müll. Arg.: A potential Source for natural antimicrobial agents. The result show that methanol extract had significant antibacterial activity against S. pneumoniae (29 mm) with minimum inhibitory concentration and maximum bacterial concentration values of 15.62 g/mL respectively. After that Kalaycıoğlu et al. 38 has studied the antimicrobial properties of chitosan films inserted with turmeric extract. Chitosan has already shown to have strong antimicrobial properties. They have found that during the 3h exposure duration when compared to the control film, chitosan films showed a significant reduction ( P < 0.05) in all microbe counts. Fang et al. 39 has studied to improve the Chinese sea bass ( Lateolabrax maculatus ) quality characteristics during cold storage and they used antimicrobial carvacrol in edible films prepared by flaxseed gum and sodium alginate. They discovered that carvacrol minimum inhibitory concentration (MIC) in the range of 0.125 to 0.5 mg/ml against various pathogenic bacteria. Yemiş et al. 40 has studied the antibacterial efficacy of soy edible coatings containing essential oils of thyme and oregano on pathogenic microorganisms in beef. They have found that soy edible coatings were found to have antibacterial efficacy against S. aureus (23 mm), L. monocytogenes and E. coli O157:H7 both with 16 mm inhibition zone diameter. 3.2. Beef Patty coated with anti-microbial active packaging film 3.2.1. Antioxidant activity of beef patties: Antioxidants have become increasingly popular in packaging materials as oxidation has become a major issue influencing food quality 41 . The results regarding antioxidant of edible film depicted highly significant effect of treatments and significant for days of storage period while their combined effect was non-significant. The results pertaining to mean values for antioxidant activity of film is in Table 3 . The results show that minimum value of antioxidant activity of film (19.91%) was recorded in T 0 that was prepared without P. wightianus while the maximum mean value was (28.55%) in T 3 with 3% P. wightianus extract. The overall effect of storage days showed variation from 24.14 to 22.80% showed the slight decrease in antioxidant capacity (Table 4 ). The Moudache et al. 42 has studied the antioxidant activity of film made from olive leaf extract and cake extracts and they reported substantial antioxidant activity that may scavenge free radicals. Antioxidant capacity of olive leaves and cake extracts ranges from 0.81 to 0.349 g equivalent Trolox per g of solution. The results of current finding were in correspondence to Ma et al. 43 they have studied the curcumin's antioxidant efficacy and release kinetics from a Tara gum/polyvinyl alcohol active film. They found that as the curcumin content increased to 5%, the DPPH scavenging capacity increased from 1.81 to 35.16% like in current research antioxidant capacity increased by increasing the concentration of P. wightianus extract. After that Dou et al. 44 has studied the physical characteristics and antioxidant activity of gelatin-sodium alginate edible films containing tea polyphenols. Tea polyphenol (TP) mixed with gelatin and sodium alginate to make active edible film. This result showed that adding tea polyphenol into a gelatin and sodium alginate film solution was an efficient way to increase the film's physical characteristics and antioxidant activity. For DPPH, the films with 2.0% TP possessed the highest values of 90.62%. Likewise, Priya et al. 45 has studied the antioxidant activity of P. wightianus . By using standards, in vitro method and they found that P. wightianus methanol had a strong scavenging impact on 2, 2-diphenyl-2-picryl hydrazyl (DPPH) free radicals was 62.36%. Table 3 Effect of Treatment on beef patties coated with active packaging film Treatments Antioxidant activity(%) pH Lipid oxidation (mg MDA/kg) Moisture (%) Total Plate Count (Log CFU/g) T o 19.91 D 5.93 A 0.29 A 72.09 A 6.70 A T 1 21.85 C 5.94 A 0.23 B 73.85 A 4.89 B T 2 23.79 B 5.97 A 0.18 C 73.17 A 4.96 B T 3 28.55 A 6.07 A 0.16 D 72.34 A 4.97 B A−c Means in column with different superscripts differ substantially T 0 = CMC + PVA + CA + Glycerol + flaxseed gel T 1 = 1% P. wightianus + CMC + PVA + CA + Glycerol + flaxseed gel T 2 = 2% P. wightianus + CMC + PVA + CA + Glycerol + flaxseed gel T 3 = 3% P. wightianus + CMC + PVA + CA + Glycerol + flaxseed gel Table 4 Effect of storage on active packaging film Storage Period Days Parameters 0 10 20 30 Antioxidant activity (%) 24.14 A 23.83 A 23.32 AB 22.80 B pH 5.92 AB 5.93 AB 6.02 AB 6.11 A Lipid oxidation (mg MDA/kg) 0.17 D 0.20 C 0.22 B 0.26 A Moisture (%) 74.53 A 73.70 A 72.56 AB 70.66 B Total Plate Count (Log CFU/g) 4.81 C 5.36 B 5.55 B 5.79 A A−D Means in column with different superscripts differ substantially 3.2.2 pH measurement of beef patties The texture of beef degrades over time as a result of the enzymatic activity of the microorganism present, which is accompanied by the dissociation of protein constituents and the synthesis of nitrogen compounds, raising the meat's pH value 46 . The results regarding pH of edible film showed non-significant ( P > 0.05 ) effect of treatments and the effect of treatments and storage on each other. While the effect of storage days showed non- significant effect on film. The mean values for pH of film is given in Table 3 show that minimum value of pH 5.93 was recorded in T 0 and the maximum mean value was 6.07 in T 3 with 3% P. wightianus extract. The overall effect of storage days showed non- significant variation in between (5.92 to 6.11) during days of storage (Table 4 ). The results of recent findings were in line previous study 27 and they have reported the effect of high alkaline pH on prevalence of Pale, Soft and Exudative (PSE) chicken protein on the formation of edible film. They have found that the film samples had better mechanical properties at pH 11.5 than the other pH-adjusted protein solutions. After that a study analyzed the shelf life of beef that can be extended using a chitosan starch film with peel extract of pomegranate and essential oil of Thymus kotschyanus 28 . They have found that the pH values of meat samples changed during storage, with the control and treated film made of 1% peel extract and 2% essential oil the highest (6.65) and lowest (5.68) pH values, respectively. After that Linghu et al. 47 has studied the effects of amino acids on production of heterocyclic amines (HCA) and their physicochemical properties in pan-fried beef patties. They have found that pH ranges from 6.41 and 6.94 in comparison to untreated samples. 3.2.3. Peroxidation of lipid in beef patties Lipid oxidation is a key cause of meat and meat product quality deterioration 48 . The results regarding lipid oxidation of edible film showed highly significant ( P < 0.01 ) effect of treatments while the effect of storage days was significant ( P < 0.05 ). The overall effect of different treatments showed variation in between 0.29 to 0.16mg/Kg (Table 3 ) and during 15 days of storage the lipid oxidation increased from 0.17 to 0.26 mg/Kg (Table 4 ). The results of our findings were in correspondence with Ouerfelli et al. 49 who have studied the effect of neem ( Azadirachta indica L .) on lipid oxidation in uncooked chilled beef patties. They have found that TBARS value range from 0.0 to 2.4mg MDA/Kg. Similarly, Fruet et al. 50 has studied the various antioxidants effects on the quality of beef patties from steers fed low-moisture distillers grain. They have found that TBARS value range from 0.2 to 0.8 mg MDA/Kg. So, we can say that P. wightianus also have antioxidant potential to reduce lipid oxidation in beef patties. 3.2.4 Moisture content of beef patties Meat, with its high moisture and nitrogen content, proper pH and fermentable carbohydrates, is an ideal medium for the proliferation of various food spoilage and poisoning bacterial and fungus species. Water is a fundamental component of most foods, and changes in moisture level can cause significant changes in food stability and quality 46 . The results regarding moisture of edible film showed non-significant ( P > 0.05 ) effect of treatments and the effect of treatments and storage on each other. However, the effect of storage days showed significant ( P < 0.05 ) effect on film. The mean values for moisture of film are given in Table 3 . The overall effect of treatment showed slight variation in between (72.09 to 72.34%) while during 30 days of storage moisture decreases from 74.53 to 70.66%. The findings of current research were similar to previous study 51 The have studied on the effect of adding edible mushroom flours ( Agaricus bisporus and Pleurotus ostreatus ) to cold-stored beef patties on physicochemical and sensory qualities. They have found that moisture content ranges from 68.78 to 70.58%. Bojorges et al. 52 has studied application of an edible film on meat containing turmeric ( Curcuma longa L). They have found that moisture content in edible film was 23.83%. Generally moisture content in meat were reported by Hammad et al. 53 they have studied the effects of freezing and refreezing on the chemical composition of beef over a 4.5-month storage period. They have found that moisture content ranges from 71.38% in beef and 75.03% in poultry. They also reported that overall moisture content was decrease with increase in storage period and it was correspondence to the findings of recent research. 3.2.5. Total Plate count (TPC)/ Total Viable Count (TVC) of beef Patties Bacterial contamination and lipid oxidation in beef during processing and storage are major causes of food borne illness and reduced shelf life 54 . The results regarding total viable count of beef patties of edible film showed a highly significant ( P < 0.01 ) effect of treatments, and storage days. The effect of treatments showed a variation in between (6.70 to 4.97 log CFU/g) and the effect of 15 days of storage showed an increase in microbial load from 4.82 to 5.80 log CFU/g. The results of recent findings were in line with Shin et al. 55 they have studied the control of microbial growth and lipid oxidation on beef products using an apple peel-based edible coating treatment. They found that after 10 days of storage, the microbiological load of the beef count of the control (uncoated beef patty) increased from 4.8 to 5.5 log CFU/g and in apple, peel-based edible coating material it decreased from 4.4 to 3.3log CFU/g during storage of 10 days. Likewise, 56 has studied the incorporation of spice essential oils into a poly-lactic acid film matrix intending to extend the microbiological and sensorial shelf life of ground beef. They reported that the overall storage period showed an increase in microbial growth of all treatments. They found that total bacterial count was increased from 4.2 to 8 log CFU/g. 3.2.6. Sensory evaluation Sensory evaluation is an experimental approach for inducing, measuring, evaluating, and construing sensory reactions for a food product. The results regarding flavor, juiciness, odor of beef patties showed highly significant ( P < 0.01 ) effect of treatment while the effect of storage days showed non-significant effect on film. The overall effect of storage days showed variation in between 7.77 to 7.55 while the treatment variations were such as 7.37 to 8.23 during 15 days of storage (Fig. 4 ). The overall effect of storage days for juiciness of patties showed non- significant ( P > 0.05 ) variation in between 7.65 to 8.27 during 15 days of storage (Fig. 4 ). The mean values for odor of beef patties showed that minimum value for odor of beef patties (6.0) recorded in T 1 that was prepared with 1% P. wightianus at 0 day of storage while the maximum mean value was 8.16 in T 3 with 3% P. wightianus extract at 10th day of storage (Fig. 3 ). The overall effect of storage days showed variation in between 6.77 to 7.55 and treatment variation were 7.88 to 8.33 during 15 days of storage (Fig. 5 ). The results show that minimum value for tenderness of beef patties 6.37 noted in T 0 at 15th day of storage while the maximum mean value was 8.96 in T 2 with 2% P. wightianus extract at 15th day of storage. The overall effect of storage days showed variation in between 8.03 to 7.89 and treatment effect varied from 7.64 to 8.33. The results for texture showed that minimum value of tenderness of beef patties 7.34 recorded in T 0 at 15th day of storage while the maximum mean value was 9.51 in T 2 with 2% P. wightianus extract at 10 day of storage (Fig. 3 ). The overall effect of storage days showed significant variation in between 7.96to 9.07 during 15 days of storage. The results regarding overall acceptability showed that minimum value 6.67 recorded in T 0 at 0 day of storage while the maximum mean value was 9.00 in T 0 at 0 day of storage. The overall effect of storage days showed significant variation in between 7.88 to 8.33 during 15 days of storage (Fig. 4 ). 4. Conclusion When compared to commercially available polymers, bio-packaging films have weak mechanical characteristics and high-water solubility and permeability. Active packaging is also becoming more important as consumers seek minimally processed and natural products. Hence, the result of this study indicated that antimicrobial edible active packaging film developed with Carboxy methylcellulose, poly vinyl alcohol, citric acid, glycerol, flaxseed gel and Phyllanthus wightianus enhanced the shelf life of beef patties. Moreover, among the treatments antimicrobial perspective of film with higher concentration of Phyllanthus wightianus was higher as in T 3 that was with 3% extract concentration. Therefore, utilization of natural antimicrobial sources as preservation of meat is a valuable replacement of synthetic chemicals. Declarations Acknowledgement: This research was supported by Guangzhou Philosophy and Social Science Development "14th Five-Year Plan" 2022 Joint Project "Research on Industrial Upgrading of Guangzhou High-end Equipment Manufacturing Industry Based on Dual Value Chain Perspective" (Project No.: 2022GZGJ79); and Institute of Food Science and Technology, Faculty of Food, Health Science and Technology, Khwaja Fareed University Engineering and Information Technology, Rahim Yar, Pakistan. Author Contributions AT, SA and KX contributed equally, they designed the study and wrote the manuscript’s first draft. MFGC and MAF conducted sample selection and data management. AL and TM, managed the literature searches and analyses. AK, TM and SZ edited the manuscript and supervised the work. ST and SA has interpreted the results and written the manuscript HL, AR and AS help in manuscript reviewing. All authors contributed to and have approved the final manuscript. Additional information We confirmed that study was in accordance with relevant institutional, national, and international guidelines and legislation Data Availability Statement The original contributions presented in the study inquiries can be directed to the corresponding author/s. Conflicts of Interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. References Robertson, G. L. Food packaging: principles and practice . (CRC press, 2016). Paunonen, S., Pitkänen, M., Vähä-Nissi, M., Leminen, V. & Kainusalmi, M. J. J. o. A. P. R. Suitability of Active and Intelligent Packaging for Local and Organic Food–A Case Study in Southern Finland. 10, 2 (2018). Abdollahzadeh, E., Nematollahi, A., Hosseini, H. J. T. i. F. S. & Technology. Composition of antimicrobial edible films and methods for assessing their antimicrobial activity: A review. 110, 291–303 (2021). 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Incorporation of spice essential oils into poly-lactic acid film matrix with the aim of extending microbiological and sensorial shelf life of ground beef. LWT 96, 482–490, doi: https://doi.org/10.1016/j.lwt.2018.05.067 (2018). Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-3913167","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":272618089,"identity":"8e2ef4a3-4196-4730-af69-465f1feb0806","order_by":0,"name":"Kangkang Xie","email":"","orcid":"","institution":"Guangzhou College of Technology and Business","correspondingAuthor":false,"prefix":"","firstName":"Kangkang","middleName":"","lastName":"Xie","suffix":""},{"id":272618090,"identity":"9fd4e943-8720-4837-a1a8-6402aaa1cc1e","order_by":1,"name":"Samreen 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on Sensory Evaluation of Beef Patty coated with active packaging film\u003c/p\u003e","description":"","filename":"Onlinedrawingimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-3913167/v1/3cb4b187ca4da569fdf52b29.png"},{"id":55265579,"identity":"b28261b9-8013-4637-8e46-81aba17d7502","added_by":"auto","created_at":"2024-04-25 02:08:33","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1301006,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-3913167/v1/f642d943-1bc8-470e-84a4-58c9d2e24a71.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Formulation and Evaluation of Phyllanthus Wightianus-Based Active Packaging Film for Meat Preservation","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eThe increased consumer awareness of synthetic packaging for food products has shifted the attention of researchers toward developing advanced eco-friendly packaging. Many concepts have been introduced in packaging, and \"Active Packaging\" (AP) is one of those. Active packaging modifies its physical state to improve its safety, sensorial, or shelf-life characteristics while preserving the food's quality. Intelligent food packaging, also known as \"active packaging,\" functions as an additional purpose above conventional packaging \u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. Active packaging enhances the product's shelf life by interacting with the consumables inside the packaging. It has been classified into systems that either absorb undesired substances or release active substances. It covers a wide range of technologies, some of which, like oxygen scavengers and moisture absorbers, have a well-defined role in the excellent consumer market, while others, such as antibacterial materials, are still in the early stages of commercialization \u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eActive packaging gently releases the added active ingredients into the food product while absorbing harmful elements like moisture, oxygen, and free radicals to preserve the quality attributes of food \u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e. The area of AP is growing substantially due to continuous changes in contemporary consumer demand and industry trends. The primary AP systems include those that produce carbon dioxide and ethanol, absorb and regulate moisture, scavenge oxygen, and work with migrating and non-migrating antimicrobial (AM) systems \u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e. Edible packaging films with antimicrobial properties have generated a lot of scientific and technical attention recently because they have the ability to improve the safety and quality of food. There are two terms, \"film\" and \"coating\" used interchangeably for packaging systems however, film is defined as a stand-alone wrapping material, whereas a coating is a thin layer that is created immediately on the surface of food and has a thickness of less than 0.3 mm \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e. Antimicrobial active packaging film can be developed using antimicrobial substances, including organic acids, bacteriocins \u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e, and extracts \u003csup\u003e\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e. The AM agents inhibit by changing the microflora's cell wall composition and showing \u0026ldquo;cidal\u0026rdquo; or \u0026ldquo;static\u0026rdquo;. These compounds ordinarily include natural antimicrobials, antioxidants, antimicrobial polymer, and biotechnology products \u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e is monoicous, sub-shrub up to 1m in loose whorls, and piliferous. Leaves alternate, distiches, oval, green above. All parts of the plant (\u003cem\u003eP. wightianus\u003c/em\u003e) have been utilized for a long time as part of ethno- medicine, as an antidiarrheal, against jaundice, and to treat dieresis. It demonstrates many biological characteristics, including \u0026ldquo;antimicrobial\u0026rdquo;, pesticide, analgesics, tissue repair, and antioxidant capabilities.\u003c/p\u003e \u003cp\u003eFlaxseed (Linumusitatissimum L.) is an individual from the variety Linum in the family Linaceae. Flaxseed is also named as functioning or bioactive food because it contains different supplements, such as bioactive compounds and alpha-linolenic acid \u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u003c/sup\u003e. In recent years, flaxseed (linseed) (\u003cem\u003eLinum usitatissimum\u003c/em\u003e) has become a popular functional food ingredient. Flaxseed gel is a type of hydrocolloid widely used in the food sector. Increased neutral polysaccharide content, such as xylose, has been demonstrated to improve flaxseed gel rheological qualities by improving shear thinning and gelling properties. Because of its favorable hydrophilicity and remissibility, this gel has many applications in meat products \u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e. The beef has a shelf life of around 3\u0026ndash;5 days at 4 \u0026ordm;C. The high quantity of moisture, oxygen, and enzyme ease the lipid oxidation, and the high protein content prevents microbial growth during shelf life \u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e. The current research emphasizes on shelf life enhancement of beef patties by using active edible packaging film. This film was prepared by using the antimicrobial properties of \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e and gelling features of flax seed as a core ingredient for its functional properties and then to determine the physicochemical, antimicrobial, textural, and sensory properties of edible active packaging film and beef patties.\u003c/p\u003e"},{"header":"2. Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 Materials\u003c/h2\u003e \u003cp\u003eAll the substances, \u003cem\u003ei.e\u003c/em\u003e., Carboxy methylcellulose, polyvinyl alcohol, citric acid, glycerol, and Mueller Hinton agar, were purchased from Sigma Aldrich and Merck Company. \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e and flaxseed were purchased from the local market of the city of Rahim Yar Khan, Punjab, Pakistan.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2 Preparation of flaxseed gel\u003c/h2\u003e \u003cp\u003eThe flaxseeds were cleaned manually by removing any foreign material, then washed and soaked with distilled water. A magnetic stirrer was used to stir the flaxseed for 5 hours. Then the water mixture was placed in a water bath at 60\u0026deg;C at 300 rpm, filtered through muslin cloth, and assembled with minimal stirring using a glass rod \u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3 Preparation of \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e extract\u003c/h2\u003e \u003cp\u003eThe dried leaves of \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e were used in a Soxhlet apparatus; the leave powder of \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e was taken in a thimble, and solvent hexane was used for extract extraction for 4 hours that may contain \u003cem\u003ee.g.\u003c/em\u003e, Caffeic acid, catechol, and pyrogallol. The extract was filtered with Whatman filter paper to get a refined extract and placed in a rotary evaporator (Daihan scientific) at 35 \u0026ordm;C during the working evaporator bath temperature was 53\u0026ordm;C, and the chiller temperature was 4\u0026ordm;C for 1 hour \u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e. The details are given in Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e1\u003c/span\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4 Active packaging film preparation\u003c/h2\u003e \u003cp\u003eThe film for active packaging was prepared by mixing Polyvinyl alcohol (5%) and carboxymethylcellulose (1%) in hot distilled water at 60\u0026ordm;C by constant stirring in the water bath (WB-22). Then glycerol (0.5%) was added as a plasticizer in a polyvinyl alcohol and carboxy methylcellulose solution with continuous stirring. After that, citric acid (5%) and flaxseed gel were added by following extract of \u003cem\u003eP. wightianus\u003c/em\u003e (1%) was added in T\u003csub\u003e1\u003c/sub\u003e, (2%) in T\u003csub\u003e2,\u003c/sub\u003e and (3%) was added in T\u003csub\u003e3\u003c/sub\u003e solution. A homogenizer was used to homogenize the solutions independently (HG-15D) for 5 minutes at 700 rpm. The prepared solution was transferred into each petri plate (15 mL) and placed in an incubator (model No. 30-1060) at 50\u0026ordm;C and 50% relative humidity for 24 hours. After drying, the intact films were carefully removed as a peel with the help of a spatula \u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e. The details are provided in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e2\u003c/span\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e2.5 Characterization of antimicrobial active packaging film\u003c/h2\u003e \u003cdiv id=\"Sec8\" class=\"Section3\"\u003e \u003ch2\u003e2.5.1 Thickness\u003c/h2\u003e \u003cp\u003eThe film thickness was measured with a handheld micrometer cut into rectangular strips of 1\u0026times; 5 cm \u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section3\"\u003e \u003ch2\u003e2.5.2 Film opacity\u003c/h2\u003e \u003cp\u003eThe film opacity was calculated using a colorimeter calibrated by using white and black plates. The film's absorbance was evaluated using a UV spectrophotometer at a wavelength of 600nm.\u003c/p\u003e \u003cp\u003e \u003cspan class=\"InlineEquation\"\u003e \u003cspan class=\"mathinline\"\u003e\\(\\text{O}\\text{P} \\left(\\text{%}\\right) = \\frac{OPb}{opw } \\times 1\\)\u003c/span\u003e \u003c/span\u003e00\u003c/p\u003e \u003cp\u003eO\u003csub\u003epb\u003c/sub\u003e and O\u003csub\u003epw\u003c/sub\u003e are opacity on at black and white background, respectively \u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section3\"\u003e \u003ch2\u003e2.5.3 Degree of swelling\u003c/h2\u003e \u003cp\u003eThe film was cut into 2\u0026times;2 pieces to assess swelling, solubility, and degree. The pieces dried at 105 \u0026ordm;C to a constant weight (M\u003csub\u003e1\u003c/sub\u003e) to achieve initial dry mass.\u003c/p\u003e \u003cp\u003eAfter weighing, the film (M\u003csub\u003e1\u003c/sub\u003e) was put into 50 ml beakers with 30 ml distilled water and held at 25\u0026ordm;C for 24 hours. The wet films were dried with filter papers before being weight (M).\u003c/p\u003e \u003cp\u003eThe swelling degree is calculated by using this equation\u003cdiv id=\"Equa\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equa\" name=\"EquationSource\"\u003e\n$$\\varvec{F}\\varvec{i}\\varvec{l}\\varvec{m} \\varvec{s}\\varvec{w}\\varvec{e}\\varvec{l}\\varvec{l}\\varvec{i}\\varvec{n}\\varvec{g} \\varvec{D}\\varvec{e}\\varvec{g}\\varvec{r}\\varvec{e}\\varvec{e}=\\frac{\\varvec{M}2-\\varvec{M}1}{\\varvec{M}1}\\varvec{x} 100$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eWhere M1\u0026thinsp;=\u0026thinsp;Initial dry mass\u003c/p\u003e \u003cp\u003eM2\u0026thinsp;=\u0026thinsp;Final dry mass \u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec11\" class=\"Section3\"\u003e \u003ch2\u003e2.5.4 Water vapor permeability\u003c/h2\u003e \u003cp\u003eEvery film sample (4cm \u0026times;4cm) used to seal test tubes containing 10g of silica (0 percent RH). All test tubes were measure and immerse in a desiccator containing distilled water. The test tubes were tested again after 24 hours to see how much they had grown in weigh \u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section3\"\u003e \u003ch2\u003e2.5.5. Antimicrobial activity\u003c/h2\u003e \u003cp\u003eDisc zone of inhibition assays were carried out as a qualitative test for antibacterial activity. The films were cut into 10 mm discs aseptically and then placed on Muller Hinton plates for \u003cem\u003eE. coli\u003c/em\u003e O157:H7 and \u003cem\u003eStaphylococcus aureus\u003c/em\u003e, which had previously been inoculated with 0.1 mL of 10\u003csup\u003e5\u003c/sup\u003e CFUs/mL inoculum. Samples were incubated for 48 h at 37\u0026deg;C. The thickness (millimeters) of the inhibition zone around the film disk (colony-free perimeter) was measured with a millimeter scale, and the growth below the film disks (the contact area of edible film with agar surface) was visually examined \u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section3\"\u003e \u003ch2\u003e2.5.6. Antioxidant activity of antimicrobial active packaging film\u003c/h2\u003e \u003cp\u003eA different volume of film sample was individually reacted at room temperature for 1 hour in the dark with 4 mL of 100mol/L DPPH methanol solution. At 517nm, the absorbance of the reaction solution was measured. The absorbance of the reaction solution and the blank are A\u003csub\u003e0\u003c/sub\u003e and A\u003csub\u003e1\u003c/sub\u003e\u003csup\u003e19\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eDPPH radical scavenging activity (%) =\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(A0-\\frac{A1}{A0}\\times 100\\)\u003c/span\u003e\u003c/span\u003e\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e2.6 Analysis of Beef Patties coated with antimicrobial active packaging film\u003c/h2\u003e \u003cdiv id=\"Sec15\" class=\"Section3\"\u003e \u003ch2\u003e2.6.1 pH measurement\u003c/h2\u003e \u003cp\u003eA hand-held pH meter was used to calculate the pH of 10% of the homogenate sample after microbiological analysis \u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section3\"\u003e \u003ch2\u003e2.6.2 Peroxidation of Lipid\u003c/h2\u003e \u003cp\u003eLipid samples were dissolved in a 30 mL mixture and glacial acetic acid (3:2, v/v). A saturated KI (1 mL) solution was added. The mixture was hand-shaken for 30 seconds before being held in the dark for another 5 minutes. The mixture was titrated against sodium thiosulphate (0.1 mol/L) after adding 100 mL distilled water until the yellow color almost vanished. After that, 1 mL of starch indicator solution (0.05%) was added. The titration was continued until the blue color completely vanished \u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section3\"\u003e \u003ch2\u003e2.6.3 Moisture content\u003c/h2\u003e \u003cp\u003eEach square film sample of 2cm\u0026times;2cm was weighed for initial weight and dried at 105\u0026ordm;C in a drying oven for 24hrs \u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec18\" class=\"Section3\"\u003e \u003ch2\u003e2.6.4 Total Plate Count (TPC) / Total viable count (TVC)\u003c/h2\u003e \u003cp\u003eThe beef patties (10g meat) were removed aseptically and homogenized in a stomacher in sterile saline. Serial dilutions were performed, with sufficient dilutions on plate count agar (PCA) for total viable count (TVC). Counts were done at 5 days interval of chilled storage and represented as log cfu/g \u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e,\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section3\"\u003e \u003ch2\u003e2.6.5 Sensory evaluation\u003c/h2\u003e \u003cp\u003eOverall acceptability on the 9-point hedonic scale of beef patties was carried out to assess the acceptability of coating by a panel of 6 judges. The panels of judges were trained and familiar with products attribute. Scales of hedonic, where 9 is equal to extremely liked and 1 is equal to extremely disliked, were used. Water was provided for rinsing mouth after each sample\u003csup\u003e\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e,\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec20\" class=\"Section3\"\u003e \u003ch2\u003e2.6.7 Statistical evaluation\u003c/h2\u003e \u003cp\u003eThe information collected for every variable was subjected to statistical evaluation in order to measure significance level \u003csup\u003e\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e. Two-factor factorial design under CRD was performed to determine results for all parameters by using statistics 8.1 software.\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e"},{"header":"3. Results and Discussion","content":"\u003cp\u003eCurrent research aimed to prepare antimicrobial edible active packaging film for beef patties by using \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e as an antimicrobial agent and flaxseed gel as a lubricating and functional ingredient.\u003c/p\u003e\n\u003cdiv id=\"Sec22\" class=\"Section2\"\u003e\n \u003ch2\u003e3.1 Characterization of anti-microbial active packaging film\u003c/h2\u003e\n \u003cdiv id=\"Sec23\" class=\"Section3\"\u003e\n \u003ch2\u003e3.1.1 Thickness of anti-microbial active packaging film\u003c/h2\u003e\n \u003cp\u003eThe thickness affects the mechanical properties, water vapour permeability, barrier characteristics, and transparency of edible films. Thinner films have better optical properties but decreased mechanical properties \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e. The thickness of the film plays a vital role in the suitability of utilizing the film for food packaging because it affects both the obstacles and mechanical features of the film. The thickness of a film is determined by its composition (solid content) and its processing parameters \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e. The results regarding the thickness of edible film depicted a non-substantial (\u003cem\u003eP\u0026thinsp;\u0026gt;\u003c/em\u003e\u0026thinsp;0.05) impact of treatments and the effect of treatments and periods on each other. The overall effect of storage days showed a significant variation between (0.046 to 0.049 mm) during 30 days of storage (Table \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e\n \u003cdiv class=\"gridtable\"\u003e\u0026nbsp;\u003ctable border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv\u003eTable 1\u003c/div\u003e\n \u003cdiv\u003e\n \u003cp\u003eEffect of Treatment on active packaging film\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eTreatments\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003ethickness (mm)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eOpacity (%)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eSwelling (%)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eWater vapour permeability (gm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003epa\u003csup\u003e4\u003c/sup\u003e)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eZone of Inhibition (mm) of \u003cem\u003eStaphylococcus aureus\u003c/em\u003e\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eZone of Inhibition (mm) of \u003cem\u003eE.coli\u003c/em\u003e\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003eo\u003c/strong\u003e\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0473\u003csup\u003eAB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e26.00\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.45\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.92\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e1\u003c/strong\u003e\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0468\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e26.51\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.52\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.83\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e19.03\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16.46\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e2\u003c/strong\u003e\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0490\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e27.97 \u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.51\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.99\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e23.40\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e19.30\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e3\u003c/strong\u003e\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0485\u003csup\u003eAB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30.32\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.83\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.08\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e25.56\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e22.1\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003ctfoot\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"7\"\u003e\u003csup\u003eA\u0026minus;c\u003c/sup\u003e Means in column with different superscripts differ substantially\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"7\"\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e0\u003c/strong\u003e\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;CMC\u0026thinsp;+\u0026thinsp;PVA\u0026thinsp;+\u0026thinsp;CA\u0026thinsp;+\u0026thinsp;Glycerol\u0026thinsp;+\u0026thinsp;flaxseed gel\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"7\"\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e1\u003c/strong\u003e\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1%\u003cem\u003eP. wightianus\u003c/em\u003e\u0026thinsp;+\u0026thinsp;CMC\u0026thinsp;+\u0026thinsp;PVA\u0026thinsp;+\u0026thinsp;CA\u0026thinsp;+\u0026thinsp;Glycerol\u0026thinsp;+\u0026thinsp;flaxseed gel\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"7\"\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e2\u003c/strong\u003e\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2%\u003cem\u003eP. wightianus\u003c/em\u003e\u0026thinsp;+\u0026thinsp;CMC\u0026thinsp;+\u0026thinsp;PVA\u0026thinsp;+\u0026thinsp;CA\u0026thinsp;+\u0026thinsp;Glycerol\u0026thinsp;+\u0026thinsp;flaxseed gel\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"7\"\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e3\u003c/strong\u003e\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;3%\u003cem\u003eP. wightianus\u003c/em\u003e\u0026thinsp;+\u0026thinsp;CMC\u0026thinsp;+\u0026thinsp;PVA\u0026thinsp;+\u0026thinsp;CA\u0026thinsp;+\u0026thinsp;Glycerol\u0026thinsp;+\u0026thinsp;flaxseed gel\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tfoot\u003e\n \u003c/table\u003e\n \u003cdiv align=\"left\" class=\"colspec\"\u003e\u003cbr\u003e\u003c/div\u003e\u0026nbsp;\u003ctable id=\"Tab1\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eEffect of storage on active packaging film\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003ccolgroup cols=\"5\"\u003e\u003c/colgroup\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\" colspan=\"4\"\u003e\n \u003cp\u003eStorage Period Days\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eParameters\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eThickness (mm)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0495\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0485\u003csup\u003eAB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0473\u003csup\u003eBC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0463\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eOpacity (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e27.7\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e27.75\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e27.76\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e27.52\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eSwelling (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6.25\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.73\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.30\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.03\u003csup\u003eD\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003ewater vapour permeability (gm\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e\u0026minus;\u0026thinsp;1\u003c/strong\u003e\u003c/sup\u003e \u003cstrong\u003es\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e\u0026minus;\u0026thinsp;1\u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003epa\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e4\u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003e)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.27\u003csup\u003eD\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.90\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.66\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.99\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003ctfoot\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"5\"\u003e\u003csup\u003eA\u0026minus;D\u003c/sup\u003e Means in column with different superscripts differ substantially\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tfoot\u003e\n \u003c/table\u003e\n \u003c/div\u003e\n \u003cp\u003eThe results of our finding were in correspondence with previous study \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e, which studied the influence of extreme alkaline pH on chicken protein edible film formation and found that lower film thickness was 68.8 \u0026micro;m, because of low moisture content. As a result, the control film had a thickness of 185.2 \u0026micro;m, which was much higher than the other groups. So, it can be stated that the loss of thickness in the current study during storage could also be due to moisture loss. The results of our findings were also similar to Mehdizadeh et al. \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e who have studied to increase the shelf life of beef by a chitosan-starch film incorporating pomegranate peel extract and \u003cem\u003eThymus kotschyanus\u003c/em\u003e essential oil. They have found that film thickness in chitosan-starch varied in the range from 0.046 to 0.063 mm. After that Utami et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e studied the effects of cinnamon bark essential oil (\u003cem\u003eCinnamomum burmannii\u003c/em\u003e) on edible film characteristics and fresh beef quality. They have found that the thickness of edible films ranged from 0.114 to 0.176 mm, slightly higher than current research findings.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec24\" class=\"Section3\"\u003e\n \u003ch2\u003e3.1.2. Opacity of anti-microbial active packaging film\u003c/h2\u003e\n \u003cp\u003eFor light-sensitive food products and customer acceptability, the transparency and color of the food packaging materials are significant parameter\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\n \u003cp\u003eThe results regarding the opacity of edible film showed a non-significant (\u003cem\u003eP\u0026thinsp;\u0026gt;\u0026thinsp;0.05\u003c/em\u003e) effect of days and the effect of treatments and storage on each other. However, the effect of treatment showed a highly significant (\u003cem\u003ep\u0026thinsp;\u0026lt;\u0026thinsp;0.01\u003c/em\u003e) effect on the film, as given in Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e. The mean values for the opacity of the film are given in Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e. The results show that a minimum opacity value of 26.00% was recorded in T\u003csub\u003e0\u003c/sub\u003e. The overall effect of storage days showed non-significant variation between (26.00 to 30.32%) during 30 days of storage (Table \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e). The resultant values of recent findings\u0026apos; opacity were higher than the previous reported study \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e which studied the development of water-resistant active packaging film prepared by carboxymethyl cellulose, polyvinyl alcohol, and aloe vera. They have found that film opacity ranges from 1.35 to 2.96% unit. Moreover, Kadam et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e studied the influence of Nigella sativa seedcake polyphenolic extracts on the physicochemical qualities of chitosan-based films. They found that film opacity ranged from 3.65 to 16.75mm, which was lower than current findings. Likewise, Sun et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e has studied the preparation and characterization of antimicrobial active food packaging film based on chitosan and incorporating apple peel polyphenols. They have found that film opacity ranges from 0.71 to 4.25 (A\u0026middot;mm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e). Similarly, Rambabu et al. \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e has studied the mango leaf extract incorporated chitosan antioxidant film for active food packaging. They have found that film opacity ranges from 0.72 to 1.28 mm. For films containing various concentrations of CA, it was observed that the opacity increased with an increase in the concentration. Film opacity is an important parameter of the packaging film, as consumers prefer to be able to see the packed food items.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec25\" class=\"Section3\"\u003e\n \u003ch2\u003e3.1.3. Degree of swelling of anti-microbial active packaging film\u003c/h2\u003e\n \u003cp\u003eThe swelling capability of a film to hold water molecules defines its swelling capability. It has been reported that an increase in the concentration of citric acid usually causes decrease in the degree of swelling of films. Moreover, the swelling properties of film are inversely proportional to the degree of crosslinking, and if the crosslinking increases, then degree of swelling automatically decreases \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\n \u003cp\u003eThe results regarding degree of swelling of edible film showed highly substantial (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.01) impact on storage days and significant (\u003cem\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.05\u003c/em\u003e) effect of treatment variation however, non-significant effect of different concentrations and different storage days on each other (Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e). The results show that after control sample the minimum value of degree of swelling 5.51% recorded in T\u003csub\u003e2\u003c/sub\u003e that was prepared with 2% \u003cem\u003eP. wightianus\u003c/em\u003e while the maximum mean value was 5.83% in T\u003csub\u003e3\u003c/sub\u003e with 3% \u003cem\u003eP. wightianus\u003c/em\u003e extract. The overall effect of treatment showed increase in between 5.45 to 5.83% from T\u003csub\u003e0\u003c/sub\u003e to T\u003csub\u003e3\u003c/sub\u003e and 6.25 to 5.04% decreases during 0 to 30 days of storage.\u003c/p\u003e\n \u003cp\u003eThe degree of swelling in a recent study was higher than that (Wulandari and Yuliatmo) \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e they studied the coating effect on edible film made from a bovine split hide gelatin on the characteristics of beef meatballs. They have found that coating concentration in soluble protein ranges from 0.178 to 0.346%. However, it was lower than the previous study\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e33\u003c/span\u003e\u003c/sup\u003e, which studied the physical chitosan properties of films containing naturally occurring antioxidants. They have found that the swelling degree of the chitosan-based film ranges from 191 to 233%. Likewise, Riaz et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e34\u003c/span\u003e\u003c/sup\u003e have also studied the antimicrobial preparation and characterization of active food packaging film based on chitosan and incorporating apple peel polyphenols. They have found that swelling degree of chitosan-based apple polyphenol film ranges from 35.79 to 56.25% treatment containing chitosan- based polyphenol significantly increased the swelling degree.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec26\" class=\"Section3\"\u003e\n \u003ch2\u003e3.1.4. Water vapour permeability of anti-microbial active packaging:\u003c/h2\u003e\n \u003cp\u003eSeparating foods from ambient atmosphere vapour is one of the key functions of preservative films, which helps to avoid or delay food degradation. The water vapour permeability of the film should be kept as low as possible because it is primarily used to inhibit moisture from transferring from the food to the environment, hence extending the shelf life of food \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e34\u003c/span\u003e\u003c/sup\u003e. Water vapour permeability of films can be affected by a variety of parameters, including the chemical nature of macromolecules, crystallinity, molecular mass, orientation, and degree of cross-linking \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e35\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\n \u003cp\u003eThe results regarding water vapour permeability of edible film showed highly substantial (\u003cem\u003ep\u0026thinsp;\u0026lt;\u0026thinsp;0.01\u003c/em\u003e) effect of storage period, treatments and for the effect of treatments and storage on each other. The effect of treatment showed substantial variation in between 1.83 gm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003epa\u003csup\u003e4\u003c/sup\u003e (T\u003csub\u003e0\u003c/sub\u003e) to 2.08 gm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003epa\u003csup\u003e4\u003c/sup\u003e (T\u003csub\u003e3\u003c/sub\u003e) and among storage days, it showed variation in between 0.27 to 2.995 gm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003epa\u003csup\u003e4\u003c/sup\u003e during storage. The results of our findings were similar to previous study \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e36\u003c/span\u003e\u003c/sup\u003e who has studied on gelatin-coated paper for beef packing. They have found that water vapour permeability ranges from 0.2855 to 0.2635(g mm m-\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e h\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e kPa\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e). However, the results of our findings were lower than Riaz et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e34\u003c/span\u003e\u003c/sup\u003e they had studied the antimicrobial preparation and characterization antimicrobial active food packaging film based on chitosan and incorporating apple peel polyphenols. They have found that water vapour permeability of chitosan-based apple polyphenol film ranges from 14.65 to10.01 gm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003epa\u003csup\u003e4\u003c/sup\u003e and treatment containing chitosan- based polyphenol significantly increased the water vapour permeability. Moreover, Kadam et al. \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e has studied the influence of polyphenolic extracts of Nigella sativa seedcake on the physicochemical properties of chitosan-based film. They have found that water vapor permeability o chitosan-based film ranges from 0.207 to 0.203 gm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003epa\u003csup\u003e4\u003c/sup\u003e.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec27\" class=\"Section3\"\u003e\n \u003ch2\u003e3.1.5 Antimicrobial Activity of anti-microbial active packaging film:\u003c/h2\u003e\n \u003cp\u003eMicrobial contamination, which causes the spread of foodborne infections, is one reason that causes food modifications. As a result, regulating or preventing food contamination is crucial, and various ways have developed to achieve this goal. One of these techniques is to utilize antimicrobial active packaging, which interacts with the packaged food to extend its shelf life \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\n \u003cp\u003eThe results show maximum inhibition zone with T\u003csub\u003e3\u003c/sub\u003e treatment with 3% concentration of antimicrobial extract for S\u003cem\u003etaphylococcus aureus\u003c/em\u003e and \u003cem\u003eE. coli\u003c/em\u003e (Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e). The Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003eA and Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003eB is also depicting zone of inhibition.\u003c/p\u003e\n \u003cp\u003eNatarajan et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e has studied the \u003cem\u003eP. wightianus\u003c/em\u003e M\u0026uuml;ll. Arg.: A potential Source for natural antimicrobial agents. The result show that methanol extract had significant antibacterial activity against \u003cem\u003eS. pneumoniae\u003c/em\u003e (29 mm) with minimum inhibitory concentration and maximum bacterial concentration values of 15.62 g/mL respectively. After that Kalaycıoğlu et al. \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e38\u003c/span\u003e\u003c/sup\u003e has studied the antimicrobial properties of chitosan films inserted with turmeric extract. Chitosan has already shown to have strong antimicrobial properties. They have found that during the 3h exposure duration when compared to the control film, chitosan films showed a significant reduction (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) in all microbe counts.\u003c/p\u003e\n \u003cp\u003eFang et al. \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e39\u003c/span\u003e\u003c/sup\u003e has studied to improve the Chinese sea bass (\u003cem\u003eLateolabrax maculatus\u003c/em\u003e) quality characteristics during cold storage and they used antimicrobial carvacrol in edible films prepared by flaxseed gum and sodium alginate. They discovered that carvacrol minimum inhibitory concentration (MIC) in the range of 0.125 to 0.5 mg/ml against various pathogenic bacteria. Yemiş et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e40\u003c/span\u003e\u003c/sup\u003e has studied the antibacterial efficacy of soy edible coatings containing essential oils of thyme and oregano on pathogenic microorganisms in beef. They have found that soy edible coatings were found to have antibacterial efficacy against \u003cem\u003eS. aureus\u003c/em\u003e (23 mm), \u003cem\u003eL. monocytogenes\u003c/em\u003e and \u003cem\u003eE. coli O157:H7\u003c/em\u003e both with 16 mm inhibition zone diameter.\u003c/p\u003e\n \u003c/div\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec28\" class=\"Section2\"\u003e\n \u003ch2\u003e3.2. Beef Patty coated with anti-microbial active packaging film\u003c/h2\u003e\n \u003cdiv id=\"Sec29\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.1. Antioxidant activity of beef patties:\u003c/h2\u003e\n \u003cp\u003eAntioxidants have become increasingly popular in packaging materials as oxidation has become a major issue influencing food quality \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e41\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\n \u003cp\u003eThe results regarding antioxidant of edible film depicted highly significant effect of treatments and significant for days of storage period while their combined effect was non-significant. The results pertaining to mean values for antioxidant activity of film is in Table \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e. The results show that minimum value of antioxidant activity of film (19.91%) was recorded in T\u003csub\u003e0\u003c/sub\u003e that was prepared without \u003cem\u003eP. wightianus\u003c/em\u003e while the maximum mean value was (28.55%) in T\u003csub\u003e3\u003c/sub\u003e with 3% \u003cem\u003eP. wightianus\u003c/em\u003e extract. The overall effect of storage days showed variation from 24.14 to 22.80% showed the slight decrease in antioxidant capacity (Table \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e). The Moudache et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e42\u003c/span\u003e\u003c/sup\u003e has studied the antioxidant activity of film made from olive leaf extract and cake extracts and they reported substantial antioxidant activity that may scavenge free radicals. Antioxidant capacity of olive leaves and cake extracts ranges from 0.81 to 0.349 g equivalent Trolox per g of solution. The results of current finding were in correspondence to Ma et al. \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e43\u003c/span\u003e\u003c/sup\u003e they have studied the curcumin\u0026apos;s antioxidant efficacy and release kinetics from a Tara gum/polyvinyl alcohol active film. They found that as the curcumin content increased to 5%, the DPPH scavenging capacity increased from 1.81 to 35.16% like in current research antioxidant capacity increased by increasing the concentration of \u003cem\u003eP. wightianus\u003c/em\u003e extract. After that Dou et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e44\u003c/span\u003e\u003c/sup\u003e has studied the physical characteristics and antioxidant activity of gelatin-sodium alginate edible films containing tea polyphenols. Tea polyphenol (TP) mixed with gelatin and sodium alginate to make active edible film. This result showed that adding tea polyphenol into a gelatin and sodium alginate film solution was an efficient way to increase the film\u0026apos;s physical characteristics and antioxidant activity. For DPPH, the films with 2.0% TP possessed the highest values of 90.62%. Likewise, Priya et al. \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e45\u003c/span\u003e\u003c/sup\u003e has studied the antioxidant activity of \u003cem\u003eP. wightianus\u003c/em\u003e. By using standards, \u003cem\u003ein vitro\u003c/em\u003e method and they found that \u003cem\u003eP. wightianus\u003c/em\u003e methanol had a strong scavenging impact on 2, 2-diphenyl-2-picryl hydrazyl (DPPH) free radicals was 62.36%.\u003c/p\u003e\n \u003cdiv class=\"gridtable\"\u003e\n \u003cdiv align=\"left\" class=\"colspec\"\u003e\u003cbr\u003e\u003c/div\u003e\u0026nbsp;\u003ctable id=\"Tab3\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eEffect of Treatment on beef patties coated with active packaging film\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003ccolgroup cols=\"6\"\u003e\u003c/colgroup\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eTreatments\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eAntioxidant activity(%)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003epH\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eLipid oxidation (mg MDA/kg)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eMoisture (%)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eTotal Plate Count (Log CFU/g)\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003eo\u003c/strong\u003e\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e19.91\u003csup\u003eD\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.93\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.29\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e72.09\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6.70\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e1\u003c/strong\u003e\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e21.85\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.94\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.23\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e73.85\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4.89\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e2\u003c/strong\u003e\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e23.79\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.97\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.18\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e73.17\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4.96\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e3\u003c/strong\u003e\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e28.55\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6.07\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.16\u003csup\u003eD\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e72.34\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4.97\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003ctfoot\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"6\"\u003e\u003csup\u003eA\u0026minus;c\u003c/sup\u003e Means in column with different superscripts differ substantially\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"6\"\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e0\u003c/strong\u003e\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;CMC\u0026thinsp;+\u0026thinsp;PVA\u0026thinsp;+\u0026thinsp;CA\u0026thinsp;+\u0026thinsp;Glycerol\u0026thinsp;+\u0026thinsp;flaxseed gel\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"6\"\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e1\u003c/strong\u003e\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1%\u003cem\u003eP. wightianus\u003c/em\u003e\u0026thinsp;+\u0026thinsp;CMC\u0026thinsp;+\u0026thinsp;PVA\u0026thinsp;+\u0026thinsp;CA\u0026thinsp;+\u0026thinsp;Glycerol\u0026thinsp;+\u0026thinsp;flaxseed gel\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"6\"\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e2\u003c/strong\u003e\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2%\u003cem\u003eP. wightianus\u003c/em\u003e\u0026thinsp;+\u0026thinsp;CMC\u0026thinsp;+\u0026thinsp;PVA\u0026thinsp;+\u0026thinsp;CA\u0026thinsp;+\u0026thinsp;Glycerol\u0026thinsp;+\u0026thinsp;flaxseed gel\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"6\"\u003e\u003cstrong\u003eT\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e3\u003c/strong\u003e\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;3%\u003cem\u003eP. wightianus\u003c/em\u003e\u0026thinsp;+\u0026thinsp;CMC\u0026thinsp;+\u0026thinsp;PVA\u0026thinsp;+\u0026thinsp;CA\u0026thinsp;+\u0026thinsp;Glycerol\u0026thinsp;+\u0026thinsp;flaxseed gel\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tfoot\u003e\n \u003c/table\u003e\n \u003c/div\u003e\n \u003cdiv class=\"gridtable\"\u003e\n \u003cdiv align=\"left\" class=\"colspec\"\u003e\u003cbr\u003e\u003c/div\u003e\u0026nbsp;\u003ctable id=\"Tab4\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eEffect of storage on active packaging film\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003ccolgroup cols=\"5\"\u003e\u003c/colgroup\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\" colspan=\"4\"\u003e\n \u003cp\u003eStorage Period Days\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eParameters\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eAntioxidant activity (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24.14\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e23.83\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e23.32\u003csup\u003eAB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e22.80\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003epH\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.92 \u003csup\u003eAB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.93\u003csup\u003eAB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6.02\u003csup\u003eAB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6.11\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eLipid oxidation (mg MDA/kg)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.17\u003csup\u003eD\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.20\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.22\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.26\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eMoisture (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e74.53\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e73.70\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e72.56\u003csup\u003eAB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e70.66\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eTotal Plate Count (Log CFU/g)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4.81\u003csup\u003eC\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.36\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.55\u003csup\u003eB\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.79\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003ctfoot\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"5\"\u003e\u003csup\u003eA\u0026minus;D\u003c/sup\u003eMeans in column with different superscripts differ substantially\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tfoot\u003e\n \u003c/table\u003e\n \u003c/div\u003e\n \u003cp\u003e\u003cbr\u003e\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec30\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.2 pH measurement of beef patties\u003c/h2\u003e\n \u003cp\u003eThe texture of beef degrades over time as a result of the enzymatic activity of the microorganism present, which is accompanied by the dissociation of protein constituents and the synthesis of nitrogen compounds, raising the meat\u0026apos;s pH value \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e46\u003c/span\u003e\u003c/sup\u003e. The results regarding pH of edible film showed non-significant (\u003cem\u003eP\u0026thinsp;\u0026gt;\u0026thinsp;0.05\u003c/em\u003e) effect of treatments and the effect of treatments and storage on each other. While the effect of storage days showed non- significant effect on film. The mean values for pH of film is given in Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e show that minimum value of pH 5.93 was recorded in T\u003csub\u003e0\u003c/sub\u003e and the maximum mean value was 6.07 in T\u003csub\u003e3\u003c/sub\u003e with 3% \u003cem\u003eP. wightianus\u003c/em\u003e extract. The overall effect of storage days showed non- significant variation in between (5.92 to 6.11) during days of storage (Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e\n \u003cp\u003eThe results of recent findings were in line previous study \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e and they have reported the effect of high alkaline pH on prevalence of Pale, Soft and Exudative (PSE) chicken protein on the formation of edible film. They have found that the film samples had better mechanical properties at pH 11.5 than the other pH-adjusted protein solutions. After that a study analyzed the shelf life of beef that can be extended using a chitosan starch film with peel extract of pomegranate and essential oil of \u003cem\u003eThymus kotschyanus\u003c/em\u003e\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e. They have found that the pH values of meat samples changed during storage, with the control and treated film made of 1% peel extract and 2% essential oil the highest (6.65) and lowest (5.68) pH values, respectively. After that Linghu et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e47\u003c/span\u003e\u003c/sup\u003e has studied the effects of amino acids on production of heterocyclic amines (HCA) and their physicochemical properties in pan-fried beef patties. They have found that pH ranges from 6.41 and 6.94 in comparison to untreated samples.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec31\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.3. Peroxidation of lipid in beef patties\u003c/h2\u003e\n \u003cp\u003eLipid oxidation is a key cause of meat and meat product quality deterioration\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e48\u003c/span\u003e\u003c/sup\u003e. The results regarding lipid oxidation of edible film showed highly significant (\u003cem\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.01\u003c/em\u003e) effect of treatments while the effect of storage days was significant (\u003cem\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.05\u003c/em\u003e). The overall effect of different treatments showed variation in between 0.29 to 0.16mg/Kg (Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e) and during 15 days of storage the lipid oxidation increased from 0.17 to 0.26 mg/Kg (Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e\n \u003cp\u003eThe results of our findings were in correspondence with Ouerfelli et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e49\u003c/span\u003e\u003c/sup\u003e who have studied the effect of neem (\u003cem\u003eAzadirachta indica L\u003c/em\u003e.) on lipid oxidation in uncooked chilled beef patties. They have found that TBARS value range from 0.0 to 2.4mg MDA/Kg. Similarly, Fruet et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e50\u003c/span\u003e\u003c/sup\u003e has studied the various antioxidants effects on the quality of beef patties from steers fed low-moisture distillers grain. They have found that TBARS value range from 0.2 to 0.8 mg MDA/Kg. So, we can say that \u003cem\u003eP. wightianus\u003c/em\u003e also have antioxidant potential to reduce lipid oxidation in beef patties.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec32\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.4 Moisture content of beef patties\u003c/h2\u003e\n \u003cp\u003eMeat, with its high moisture and nitrogen content, proper pH and fermentable carbohydrates, is an ideal medium for the proliferation of various food spoilage and poisoning bacterial and fungus species. Water is a fundamental component of most foods, and changes in moisture level can cause significant changes in food stability and quality \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e46\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\n \u003cp\u003eThe results regarding moisture of edible film showed non-significant (\u003cem\u003eP\u0026thinsp;\u0026gt;\u0026thinsp;0.05\u003c/em\u003e) effect of treatments and the effect of treatments and storage on each other. However, the effect of storage days showed significant (\u003cem\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.05\u003c/em\u003e) effect on film. The mean values for moisture of film are given in Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e. The overall effect of treatment showed slight variation in between (72.09 to 72.34%) while during 30 days of storage moisture decreases from 74.53 to 70.66%.\u003c/p\u003e\n \u003cp\u003eThe findings of current research were similar to previous study \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e51\u003c/span\u003e\u003c/sup\u003e The have studied on the effect of adding edible mushroom flours (\u003cem\u003eAgaricus bisporus\u003c/em\u003e and \u003cem\u003ePleurotus ostreatus\u003c/em\u003e) to cold-stored beef patties on physicochemical and sensory qualities. They have found that moisture content ranges from 68.78 to 70.58%. Bojorges et al. \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e52\u003c/span\u003e\u003c/sup\u003e has studied application of an edible film on meat containing turmeric (\u003cem\u003eCurcuma longa\u003c/em\u003e L). They have found that moisture content in edible film was 23.83%. Generally moisture content in meat were reported by Hammad et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e53\u003c/span\u003e\u003c/sup\u003e they have studied the effects of freezing and refreezing on the chemical composition of beef over a 4.5-month storage period. They have found that moisture content ranges from 71.38% in beef and 75.03% in poultry. They also reported that overall moisture content was decrease with increase in storage period and it was correspondence to the findings of recent research.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec33\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.5. Total Plate count (TPC)/ Total Viable Count (TVC) of beef Patties\u003c/h2\u003e\n \u003cp\u003eBacterial contamination and lipid oxidation in beef during processing and storage are major causes of food borne illness and reduced shelf life \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e54\u003c/span\u003e\u003c/sup\u003e. The results regarding total viable count of beef patties of edible film showed a highly significant (\u003cem\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.01\u003c/em\u003e) effect of treatments, and storage days. The effect of treatments showed a variation in between (6.70 to 4.97 log CFU/g) and the effect of 15 days of storage showed an increase in microbial load from 4.82 to 5.80 log CFU/g.\u003c/p\u003e\n \u003cp\u003eThe results of recent findings were in line with Shin et al.\u003csup\u003e\u003cspan class=\"CitationRef\"\u003e55\u003c/span\u003e\u003c/sup\u003e they have studied the control of microbial growth and lipid oxidation on beef products using an apple peel-based edible coating treatment. They found that after 10 days of storage, the microbiological load of the beef count of the control (uncoated beef patty) increased from 4.8 to 5.5 log CFU/g and in apple, peel-based edible coating material it decreased from 4.4 to 3.3log CFU/g during storage of 10 days. Likewise, \u003csup\u003e\u003cspan class=\"CitationRef\"\u003e56\u003c/span\u003e\u003c/sup\u003e has studied the incorporation of spice essential oils into a poly-lactic acid film matrix intending to extend the microbiological and sensorial shelf life of ground beef. They reported that the overall storage period showed an increase in microbial growth of all treatments. They found that total bacterial count was increased from 4.2 to 8 log CFU/g.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec34\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.6. Sensory evaluation\u003c/h2\u003e\n \u003cp\u003eSensory evaluation is an experimental approach for inducing, measuring, evaluating, and construing sensory reactions for a food product. The results regarding flavor, juiciness, odor of beef patties showed highly significant (\u003cem\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.01\u003c/em\u003e) effect of treatment while the effect of storage days showed non-significant effect on film. The overall effect of storage days showed variation in between 7.77 to 7.55 while the treatment variations were such as 7.37 to 8.23 during 15 days of storage (Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e). The overall effect of storage days for juiciness of patties showed non- significant (\u003cem\u003eP\u0026thinsp;\u0026gt;\u0026thinsp;0.05\u003c/em\u003e) variation in between 7.65 to 8.27 during 15 days of storage (Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e). The mean values for odor of beef patties showed that minimum value for odor of beef patties (6.0) recorded in T\u003csub\u003e1\u003c/sub\u003e that was prepared with 1% \u003cem\u003eP. wightianus\u003c/em\u003e at 0 day of storage while the maximum mean value was 8.16 in T\u003csub\u003e3\u003c/sub\u003e with 3% \u003cem\u003eP. wightianus\u003c/em\u003e extract at 10th day of storage (Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e). The overall effect of storage days showed variation in between 6.77 to 7.55 and treatment variation were 7.88 to 8.33 during 15 days of storage (Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e5\u003c/span\u003e). The results show that minimum value for tenderness of beef patties 6.37 noted in T\u003csub\u003e0\u003c/sub\u003e at 15th day of storage while the maximum mean value was 8.96 in T\u003csub\u003e2\u003c/sub\u003e with 2% \u003cem\u003eP. wightianus\u003c/em\u003e extract at 15th day of storage. The overall effect of storage days showed variation in between 8.03 to 7.89 and treatment effect varied from 7.64 to 8.33. The results for texture showed that minimum value of tenderness of beef patties 7.34 recorded in T\u003csub\u003e0\u003c/sub\u003e at 15th day of storage while the maximum mean value was 9.51 in T\u003csub\u003e2\u003c/sub\u003e with 2% \u003cem\u003eP. wightianus\u003c/em\u003e extract at 10 day of storage (Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e). The overall effect of storage days showed significant variation in between 7.96to 9.07 during 15 days of storage. The results regarding overall acceptability showed that minimum value 6.67 recorded in T\u003csub\u003e0\u003c/sub\u003e at 0 day of storage while the maximum mean value was 9.00 in T\u003csub\u003e0\u003c/sub\u003e at 0 day of storage. The overall effect of storage days showed significant variation in between 7.88 to 8.33 during 15 days of storage (Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e\n \u003c/div\u003e\n\u003c/div\u003e"},{"header":"4. Conclusion","content":"\u003cp\u003eWhen compared to commercially available polymers, bio-packaging films have weak mechanical characteristics and high-water solubility and permeability. Active packaging is also becoming more important as consumers seek minimally processed and natural products. Hence, the result of this study indicated that antimicrobial edible active packaging film developed with Carboxy methylcellulose, poly vinyl alcohol, citric acid, glycerol, flaxseed gel and \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e enhanced the shelf life of beef patties. Moreover, among the treatments antimicrobial perspective of film with higher concentration of \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e was higher as in T\u003csub\u003e3\u003c/sub\u003e that was with 3% extract concentration. Therefore, utilization of natural antimicrobial sources as preservation of meat is a valuable replacement of synthetic chemicals.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgement:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research was supported by Guangzhou Philosophy and Social Science Development \u0026quot;14th Five-Year Plan\u0026quot; 2022 Joint Project \u0026quot;Research on Industrial Upgrading of Guangzhou High-end Equipment Manufacturing Industry Based on Dual Value Chain Perspective\u0026quot; (Project No.: 2022GZGJ79); and Institute of Food Science and Technology, Faculty of Food, Health Science and Technology, Khwaja Fareed University Engineering and Information Technology, Rahim Yar, Pakistan.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAT, SA and KX contributed equally, they designed the study and wrote the manuscript\u0026rsquo;s first draft. MFGC and MAF conducted sample selection and data management. AL and TM, managed the literature searches and analyses. AK, TM and SZ edited the manuscript and supervised the work. ST and SA has interpreted the results and written the manuscript HL, AR and AS help in manuscript reviewing. All authors contributed to and have approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAdditional information\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe confirmed that study was in accordance with relevant institutional, national, and international guidelines and legislation\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability Statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe original contributions presented in the study inquiries can be directed to the corresponding author/s.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflicts of Interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eRobertson, G. 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LWT 84, 183\u0026ndash;188 (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTalebi, F. \u003cem\u003eet al.\u003c/em\u003e Incorporation of spice essential oils into poly-lactic acid film matrix with the aim of extending microbiological and sensorial shelf life of ground beef. LWT 96, 482\u0026ndash;490, doi:\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/j.lwt.2018.05.067\u003c/span\u003e\u003cspan address=\"10.1016/j.lwt.2018.05.067\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2018).\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Phyllanthus wightianus, Flaxseed gel, Active film, Antimicrobial packaging film, Beef patties, Storage study","lastPublishedDoi":"10.21203/rs.3.rs-3913167/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-3913167/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eA biopolymer-based edible film was prepared using \u003cem\u003ePhyllanthus wightianus\u003c/em\u003e to extend the shelf life of beef patties. For this purpose, the film was prepared by using polyvinyl alcohol (PVA, 5% W/V), carboxymethyl cellulose (CMC, 1% W/V), glycerol (0.1% V/V) as a plasticizer, flaxseed gel and \u003cem\u003eP. wightianus\u003c/em\u003e extract @ 0, 1, 2 and 3% in various treatments. The film was analyzed at the interval of 0, 10, 20, and 30th days at 25\u0026deg;C for thickness, the film's opacity, degree of solubility and swelling, water vapor permeability, and \u003cem\u003eStaphylococcus aureus\u003c/em\u003e and \u003cem\u003eEscherichia coli\u003c/em\u003e antibacterial activity. After a month of storage study, the developed film was used to coat beef patties and analyzed for antioxidants, pH, peroxidation, and moisture. A total plate count test was performed for antimicrobial analysis, and beef patties were further evaluated for sensory evaluation parameters. The results showed that film has substantial antimicrobial potential suitable for longer storage and helpful in delaying the spoilage of beef patties by controlling lipid peroxidation and microbial growth of meat spoilage bacteria, especially \u003cem\u003eS. aureus\u003c/em\u003e and \u003cem\u003eE. coli.\u003c/em\u003e In conclusion, the films developed with 3% \u003cem\u003eP. wightianus\u003c/em\u003e extract and flaxseed gel prolong the shelf life of beef patties throughout storage.\u003c/p\u003e","manuscriptTitle":"Formulation and Evaluation of Phyllanthus Wightianus-Based Active Packaging Film for Meat Preservation","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-02-14 17:36:16","doi":"10.21203/rs.3.rs-3913167/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"a03403f8-ce79-4e54-b530-40573496950f","owner":[],"postedDate":"February 14th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":28733988,"name":"Biological sciences/Microbiology"},{"id":28733989,"name":"Physical sciences/Materials science/Techniques and instrumentation/Characterization and analytical techniques"}],"tags":[],"updatedAt":"2024-04-24T16:05:49+00:00","versionOfRecord":[],"versionCreatedAt":"2024-02-14 17:36:16","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-3913167","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-3913167","identity":"rs-3913167","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}