在提出的研究中,从海洋中分离的枯草芽孢杆菌AG4菌株进行了形态学鉴定,生理,系统发育特征和16SrRNA序列的检查。从枯草芽孢杆菌菌株中提取并分离出新型胞外多糖(EPSR4)作为胞外多糖(EPS)的主要部分。结构表征分析表明,EPSR4是β-糖苷硫酸化杂多糖(48.2%),分子量(Mw)为1.48×104g/mol,不含糖醛酸。单糖含量分析表明EPSR4由葡萄糖组成,鼠李糖和阿拉伯糖单糖的摩尔比分别为5:1:3。形态分析表明,EPSR4具有高结晶度和显著的孔隙率,其在脂质相中的聚集和构象可能对EPSR4的生物活性产生显著影响。通过研究EPSR4的抗氧化性,筛选并评价其生物活性,细胞毒性,抗炎,和抗阿尔茨海默氏症的活动。抗氧化活性结果表明,EPSR4在1500µg/mL时对DPPH自由基具有97.6%的清除活性,IC50值为300µg/mL,1500µg/mL时对过氧化氢自由基的作用为64.8%(IC50=1500µg/mL,30分钟)。此外,EPSR4对T-24(膀胱癌)的增殖表现出相当大的抑制活性,A-549(肺癌)和HepG-2(肝细胞癌)癌细胞系,IC50为244µg/mL,148µg/mL和123µg/mL,分别。对抗炎活性的评估表明,EPSR4具有有效的脂氧合酶(LOX)抑制活性(IC50为54.3µg/mL),并且对膜稳定有相当大的影响(IC50=112.2±1.2µg/mL),同时显示环氧合酶(COX2)抑制活性高达125µg/mL。最后,EPSR4对乙酰胆碱酯酶活性显示出相当大的抑制活性。一起来看,这项研究表明,枯草芽孢杆菌菌株AG4可以被认为是新型EPS的潜在天然来源,具有强大的生物活性,对医疗保健系统有用。
In the presented study, Bacillus subtilis strain AG4 isolated from marine was identified based on morphological, physiological, phylogenetic characteristics and an examination of 16S rRNA sequences. Novel exopolysaccharide (EPSR4) was extracted and isolated from the Bacillus subtilis strain as a major fraction of exopolysaccharide (EPS). The analysis of structural characterization indicated that EPSR4 is a β-glycosidic sulphated heteropolysaccharide (48.2%) with a molecular weight (Mw) of 1.48 × 104 g/mole and has no uronic acid. Analysis of monosaccharide content revealed that EPSR4 consists of glucose, rhamnose and arabinose monosaccharide in a molar ratio of 5:1:3, respectively. Morphological analysis revealed that EPSR4 possess a high crystallinity degree with a significant degree of porosity, and its aggregation and conformation in the lipid phase might have a significant impact on the bioactivity of EPSR4. The biological activity of EPSR4 was screened and evaluated by investigating its antioxidant, cytotoxicity, anti-inflammatory, and anti-Alzheimer activities. The antioxidant activity results showed that EPSR4 has 97.6% scavenging activity toward DPPH free radicals at 1500 µg/mL, with an IC50 value of 300 µg/mL, and 64.8% at 1500 µg/mL toward hydrogen peroxide free radicals (IC50 = 1500 µg/mL, 30 min). Furthermore, EPSR4 exhibited considerable inhibitory activity towards the proliferation of T-24 (bladder carcinoma), A-549 (lung cancer) and HepG-2 (hepatocellular carcinoma) cancer cell lines with IC50 of 244 µg/mL, 148 µg/mL and 123 µg/mL, respectively. An evaluation of anti-inflammatory activity revealed that EPSR4 has potent lipoxygenase (LOX) inhibitory activity (IC50 of 54.3 µg/mL) and a considerable effect on membrane stabilization (IC50 = 112.2 ± 1.2 µg/mL), while it showed cyclooxygenase (COX2) inhibitory activity up to 125 µg/mL. Finally, EPSR4 showed considerable inhibitory activity towards acetylcholine esterase activity. Taken together, this study reveals that Bacillus subtilis strain AG4 could be considered as a potential natural source of novel EPS with potent biological activities that would be useful for the healthcare system.