الفهرس 
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Abstract
Hyaluronic acid is a naturally occurring biopolymer, which serves important biological functions in humans. HA is extensively applied in both biomedical and cosmetic applications such as viscosupplementation for osteoarthritis treatment, surgical aid in eye surgery, otosclerosis surgery, non-reducing disc displacement during orthodontic treatment, scaffolding for tissue engineering, wound healing process, drug delivery agent in cancer treatment and beauty applications including antiaging, skin hydration and dermal filler as well as treatment of wrinkles. Also, the antioxidant activity of hyaluronic acid in the body acts as an efficient “scavenger” of free radicals.
Overexposure of skin to UV irradiation can cause sunburn and inflammations that can lead to skin photoaging and cancer. Therefore, the development of substances that inhibit, treat and repair harmful UV effects on skin is needed.
In the present study, eighteen bacterial isolates were isolated from three different sources (two different breast milk and kefir) on MRS, kanamycin aesculin azide and KF streptococcus agar media. Also, the strains L. plantarum strain M,
L. plantarum strain St3, L. casei 1922 and L. acidophilus 23431 were used in the study. The preliminary screening for hyaluronic
acid production by twenty-two bacterial isolates grown on skimmed milk medium was carried out. The ability of the isolates to produce HA was evaluated by turbidimetric method using CTAB reagent. Fifteen isolates were able to produce HA, while seven isolates (K1, K3, H1, H2, Z1, Z2 and Z5) were unable to produce it. The produced HA was qualitatively characterized by HPLC in comparison to the standard HA. Eight bacterial isolates (K4, K11, K33, H3, H4, H5, St3 and 1922) showed high HA productivity ranged from 3 to 4.8 mg/ml while four isolates (K2, K13, K44 and Z4) revealed moderate HA concentrations ranged from 1.9 to 2.4 mg/ml. Three isolates (Z3, M and 23431) showed mild HA concentrations 0.2, 0.5 and 0.5 mg/ml, respectively.
FTIR analysis was performed on the HA produced by the eight bacterial isolates. The FTIR spectra of the produced HA by the bacterial isolates and standard HA showed the same fingerprint. The most promising HA producing probiotic isolates K11 and St3 were selected and identified by 16S rRNA gene sequence as Enterococcus durans strain K11 (ON359827) and Lactiplantibacillus plantarum strain St3 (KY508301), respectively.
The effect of environmental conditions on the production of HA by E. durans strain K11 and L. plantarum strain St3 was examined including temperature, pH, different concentrations of
glucose (as carbon source) and yeast extract (as nitrogen source) as well as different minerals concentrations (MgSO4, MnSO4 and FeSO4). The maximum amounts of HA produced by E. durans strain K11 and L. plantarum strain St3 on skimmed milk medium and under the optimum cultivation conditions obtained from this study (incubation temperature, 37 oC; initial pH, 6; glucose concentration, 4% and without supplementation with yeast extract or mineral ions) were 5.4 ± 0.1 and 5.3 ± 0.2 mg/ml, respectively. Decreasing or increasing the temperature, pH, and glucose than optimal condition was accompanied by decrease in HA production by both strains. While it is noticeable from the results that not adding yeast extract or mineral ions is favored in the production of HA by both isolates.
The antioxidant activity of the extracted HA from both isolates was evaluated using DPPH scavenging method. The antioxidant activity of the HA produced by E. durans strain K11 and L. plantarum strain St3 were 65.4 ± 0.2% and 66.6 ± 0.1%, respectively.
The effect of HA on the damage resulted from UVB irradiation on human dermal keratinocytes was in vitro evaluated using MTT assay. The viability of the irradiated untreated cells was 76 % while the viability of irradiated pre-treated cells, with HA produced by E. durans strain K11 and L. plantarum strain
St3, was 91.3 and 91.4 %, respectively. Also, the viability of irradiated post-treated cells was 86 and 88.5% for HA produced by Enterococcus durans strain K11 and L. plantarum strain St3, respectively.