الفهرس 
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Abstract
The main purpose of the present study is the highest production, purification, characterization of hyaluronidase enzyme produced by potent local bacterial strain. Moreover, hyaluronidase was studied for anticancer activity against tumor cells, antioxidant and filler injection. The obtain results are summarized as follow
1. Twelve isolates; 11 bacterial isolates and one actinobacterial isolate were isolated from rooster comb.
2. These isolates were screened for their hyaluronidase productivity, R3 isolate was selected as the highest producer isolate for hyase.
3. The selected isolate was identified as Brucella intermedia based on the morphological, biochemical and molecular characteristics. Its 16S rRNA gene sequence was submitted to Gene Bank under accession numbers OR794010.
4. The physical factors affecting hyaluronidase production by the selected isolate were studied. Using one- factor at-a time approach, it was found that, the maximum hyase productivity was achieved using modified minimal salt medium with 1% hyaluronic acid, pH 7 and incubation for 2 days at 30ºC under shaking (150 rpm/min) condition.
5. Plackett-Burman statistical design was used to determine the most significant nutritional factors affecting enzyme activity. It was found to be CaBr and Nicotinic acid.
6. Central composite design was used to determine the optimum level of each significant factor giving the highest enzyme production. The results revealed that using CaBr (99.767mM) and Nicotinic acid (5.508 µg /l) increased enzyme production 4 times compared to the basal medium
7. The purification of hyase enzyme using 70% ammonium sulphate, DEAE sepharose and Sephacrle S100 increased the enzyme activity by 1.53, 5.2 and 9.3 folds, respectively.
8. Pure hyase enzyme had molecular weight of 65 kDa.
9. Factors affecting the activity of purified hyase was investigated;
a- The maximum hyase activity of B. intermedia strain was attained after incubation for 30 min at 37 ºC.
b- The optimum pH for maximum activity of the purified hyaluronidase was 7.
c- The purified Hyaluronidase enzyme exhibited higher thermal stability at 45 ºC for 30 minutes.
10. The purified hyaluronidase enzyme exhibited anti-cancer activity against Caco-2, A549, A431 and MDA cell lines with IC50 values of 89.34, 73.02, 61.236 and 55.66, respectively after 24h and 72.88, 54.12, 37.40 and 31.52 respectively after 72h. It also exhibited biosafety against HSF with IC50 of 310 after 24h and 276 after 72h. The outcomes demonstrated that hyase enzyme could be applied in medical and pharmaceutical applications.
11. The purified hyaluronidase from B. intermedia MEFS strain was also exhibited antioxidant activities by two in vitro assays: DPPH scavenging capacity with IC50 of 67.13 µg/mL and total antioxidant capacity with IC50 of 50.59 µg/mL.
12.The role of hyase in the treatment of dermal filler complications was also studied in hairless mice injected with hyaluronic acid dermal filler and found that complete loss of filler surface projection was obtained after 72h of hyaluronidase treatment. This finding also confirmed by histological examination.