Effect of Zinc Oxide Nanoparticles Synthesis by ( Zingiber Officinale ) Ginger Extract in Some Microbiological Quality of Fresh Chicken Meat During Referginated Storage

This study, include the green synthesis of ZnO nanoparticles using ethanol extract of ( Zingiber officinale ) ginger was investigated in order to evaluate its antibacterial effect on fresh chicken meat. To Determining the microbiological quality of meat involves performing various plate counts indicators include, total bacterial count (TBC) and the total coliform counts TCC the UV-Visible Spectrophotometry and FTiR (Fourier Transform infrared spectroscopy) were used to characterize the GnZnO nanoparticles. A total of 180 samples of fresh chicken meat were subjected to (0.5, 1, and 1.5) mg/ml of GnZnONPs and bacteriologically examined was done . significant absorption of UV region at peak370nm verified the presence of ZnO NPs. The FT-IR spectra recorded the existence of biomolecules responsible for the reduction and capping of the green synthesized , Also observed result of a meat sample treated with GnZnONPs showed a significant (p < 0.05) reduction in (TBC) through 2, 5, and 7 days of storage at 4°C (1.5 mg). While the highest count recorded in control (untreated samples) reached 11.360.04 CFU.g-1 at the end of the experiment, the TCC also decreased significantly (p< 0.05) after 5,7 days of storage at 4 0 C when the meat samples were treated with GnZnONPs compared to the control sample. The findings show that treating poultry meat with GnZNoNPs extends the shelf-life of the meat, and maintain its quality throughout the storage period.


Introduction
Foodborne illnesses attributed to the consumption of poultry meat and its processed products are a significant public health problem around the world. It can be contaminated with a variety of microbes, including those capable of deteriorating the product during refrigerated conditions as well as some foodborne pathogens (1). Microorganisms that can contaminate poultry meat and its products are mainly transferred to humans through eating raw or undercooked chicken meat (2). Several studies have shown that chicken meat consumption is associated with the incidence of outbreaks of foodborne diseases (3). Chicken meat is the most preferred meat because it is an economical and available source of protein with a low fat content and low in cholesterol compared to other meats. , skin of raw chicken meat and cuts directly contact the air and equipment surface, so it is easily contaminated.
(4) In order to reduce the economic burden of spoilage of raw poultry meat, methods for extending the shelf life and overall safety/quality of the meat are being researched. This is important because the initial bacterial load in the meat is the primary limitation on shelf life and overall safety/quality of the meat (5). When raw meat is kept refrigerated, it can spoil in two ways: through oxidative rancidity and through microbial growth.
(6). Antimicrobial preservatives are ingredients that are used to extend the shelf life of meat by reducing the growth of microorganisms during slaughter, transportation, processing, and storage. Nanoparticles are employed in a variety of ways. Metal oxide nanoparticles are mainly used to reduce food pathogens and prevent food contamination. Nanoparticles having a large surface area and the ability to change their size can be employed in a variety of applications. In the food industry, nanoparticles help to enhance cleanliness, prolong product shelf life, and avoid food-borne illnesses and chemical contamination (7). Food processing and preservation ( nanocomposites, nanocoatings, nanosensors, edible coating NPs, etc.) and food packaging (nanocoatings, nanosensors, nanocomposites, etc.) are examples of NP applications in the food industry ( 8 ). Plant extract-based green synthesis of NPs has several advantages: it is nonpathogenic, cost-effective, creates a large number of metabolites, and is environmentally friendly. Ginger (Zingiber officinale) underground rhizomes are the medicinally useful part (9 ). All Ginger's major active ingredients, such as zingerone, gingerol, zingiberene, and shogaols, are known to possess antioxidant activities Antimicrobial and antioxidant nanoparticles could be a significant development in the development of antimicrobial therapeutic agents for the prevention and reduction of food pathogenic microorganisms and deterioration ( 10). The goal of this research is to develop an alternative to synthesize and characterize zinc oxide nanoparticles (ZnONPs) using ginger extract and evaluate the antibacterial effects.to extend its shelf life of the fresh chicken meat

Green -synthesis of zinc oxide nanoparticles (ZnONPs)
Green -synthesis of ZnO nanoparticles was preferred by using ethanolic (Zingiber officinale) ginger root extracts 70% as studies by (11). Deionized water was used to prepare 50 mL of 0.01 M Zinc acetate dehydrate (Sigma Aldrich, Germany ) . In a magnetic stirrer, 500 ml of extract solution was slowly added by continuous stirring. 1.0 M sodium hydroxide( Fluka , Germany) was used to maintain pH at 12 , the mixture was stirred for 2 hours until a white precipitate formed, after which it was centrifuged for 10 minutes at 10,000rpm. Pellets were washed in deionized water and dried overnight in a hot air oven at 200°C. The resulting white powder was carefully collected and sent to be characterized. UV spectrum (Schimadzu 1601 spectrophotometer) in 250-900 nm range and Vol. Fourier-transform infrared (FTIR) Shemadzu (Germany) spectrum, to performed the synthesized NPs. By adding a specific quantity of the organic solvent dimethyl sulfoxide (DMSO) to the solution a measured volume of ZnO nanoparticles (0.5,1, and 1.5 g/mL), ZnO nanoparticles were prepared., by allowed the mixture to sonicate for a few minutes 2. Collection and preparation of of meat samples: A total of 180 fresh meat samples (500 g ) were collected from a local abattoir in Al Diwaniyah city and transported in ice boxes directly to the vetrenary college / public health laboratory. GZno-NPs powder dissolved in solution at the following concentrations (0.5mg/ml, 1mg/ml, and 1.5mg/ml) in deionized water the fresh meat samples divided in Four groups in accordance with ( Elsaid et al., 2019) First, the control group with out treatment. Meat sample dipping in nanomaterial . for I hr at room temperature (25 °C). then drained for 2 minutes before being packaged in polyethylene bags, labeled, and stored at 4 °C in order to evaluate microbiological count TBC Total Bacterial Counts And Total Coliforms Counts (TCC ) during 0, 2, 5, and 7 days of refrigerated storage.

3.Preparation of samples for microbiological analysis :
The samples were prepared in accordance with FDA guidelines for bacteriological analysis (2001). homogenize 25 g of each sample with 225 mL of BPW ) Himedia / India ( solution . The total bacterial count was determined by preparing decimal serial dilutions (10-1 to 10-6) of each meat sample in sterile 0.1 % (wt/v) peptone water and then pouring onto plates of nutritional agar at 45°C, for 24 hr. each dilution was done in 3 triplicate. While total count of coliforms was estimated by serial dilutions (10-1 to 10-6) of each meat sample in sterile peptone water 0.1. Pour plated in violet red bile agar ( Himedia / India) for each dilution incubated for 48hr then Multiplication of the average number of bacterial colonies by reciprocal dilution (cfu/ml).

Statistical analysis:
The significance of the differences in mean values has to be determined. As a result of this research, two-way ANOVA was used to statistically evaluate the acquired findings using the SPSS program (SPSS 19.0, Chicago, IL, USA). The post-hoc Duncan's test was used. The data was provided as Mean ±Standard Error (SE) (13).

Gn-ZnONPs Characterization :
a-Fourier-transform infrared Shemadzu (Germany) spectrum (FTIR): was used to investigated the functional groups of biomolecules in the ginger extract which are responsible for ZnO nanoparticle reduction and stabilization. Figure (1) The peaks at (3430) cm 1, (415) cm1, 1516 cm 1, 1384 cm 1, and 872 cm-1 correspond to N-H(Amine) or amide groups, Zn-O bond, and C=O stretch, indicating ketones. Respectively. (14) ( 15) When making Zn2 oxid nanoparticles, it is preferable to use natural stabilizers instead of artificial or chemical ones., Nanoparticlesize,shape, and morphology are significantly influenced by the phytochemicals in the plant extract. Consequently, the nanoparticles are extremely stable and have a narrow particle size distribution (16 ) . b-UV-visible absorption spectrum : The synthesis of ZnO NPs was confirmed by( Schimadzu 1601 spectrophotometer) in 250-900 nm range a significant absorption band with a maximum wavelength at 370 nm was indicated (fig 2). This sharp peak confirm that the particles are in Nano sized, according to the spectrum which is identical to results has been reported for ZnO NPs (17).   (1) and figure (3 ) The initial mean of total bacterial count for ( control group) untreated meat samples was observed to be 3.95±0.02 CFU.g-1 these value was increased significantly (P ≤ 0.05) during day of storage, the result showed significantly highest count recorded in control (untreated samples ) reaching 7.93±0.02 CFU.g-1 and 11.36±0.04 CFU.g-1 after storage for 5 day and 7 day respectively. While the sample treated with GnZnONPs shown a significant (p≤ 0.05). reduction on total bacterial count after 2, 5, and 7 days of storage at 4°C., it was observed the mean of TBC was the lowest recorded in sample treated with T3( 1.5 mg) 5.08±0.01 CFU.g-1 At 7 day 0f storage although there was decreases in TBC in sample treated with T1 7.28±0.01 CFU.g-1. And T2 6.84±0.05 CFU.g-1. In7 days of storage While The effect of storage periods on all groups was significant (P<0.05), the count increase gradually with the increase of storage periods, the highest count recorded in the last periods at7 days of storage in all groups, and the lowest count recorded in the first periods zero days of storage in all groups. However, there is a significant difference (P<0.05) between T1 ,T2 and T3 through the storage periods the highest significantly decrease (p<0.05) in sample treated with T3 observed result of the total bacterial counts after 7 days of refrigerated storage at 4 0C were affected significantly (p< 0.05) by treated the meat samples GnZnONPs compared with control . nanoparticles play important role in bacterial inhibition in samples treated with ZnONPs compared with samples without treated , these results agreed with mentioned by (19 ) that indicated during a study bio synthesis ZnO NPs exhibited strong antimicrobial activity against food-borne bacteria. also maintained the organoleptic features of chicken fillet during storage Also, agreement with mentioned by (20) (25), (26) and (27) when uses ZNONPs as antibacterial growth , FDA( Food and Drug Administration) approved zinc compounds can be used in food production because they have been presumed safe for human consumption by the organization (28) . Further study in this area is needed in the future.

Conclusion
Nanotechnology is a novel application that has been deployed in the meat production . The antibacterial effect of green synthesis ZnO-NPs is concentration dependent . Bacterial count was considerably (P< 0.05) lower in samples treated with ZnO NPs than in controls. It demonstrates that ZnO NPs' has potent antibacterial action. The use of nanoparticles in the meat industry has effectively improved meat quality and safety. The US Food and Drug Administration has categorized zinc compounds as a generally recognized as safe (GRAS) substance, allowing them to be utilized in the food sector. More study should be conducted in this area in the future.