Phuong Nguyen Thi Mai, Van Quang Ngo, Mai Ta Thu, Anh Nguyen Vu, Kuhakarn Chutima, Reutrakul Vichai, Bolhuis Albert
Institute of Biotechnology, Vietnam Academy of Science and Technology, 18 Hoang Quoc Viet Road, Hanoi, Viet Nam; Graduate University of Science and Technology, Vietnam Academy of Science and Technology, 18 Hoang Quoc Viet Road, Hanoi, Viet Nam.
Institute of Chemistry, Vietnam Academy of Science and Technology, 18 Hoang Quoc Viet Road, Hanoi, Viet Nam.
Asian Pac J Trop Med. 2017 Dec;10(12):1154-1160. doi: 10.1016/j.apjtm.2017.10.022. Epub 2017 Oct 31.
To isolate α-mangostin (AMG) from the peels of mangosteen (Garcinia mangostana L.), grown in Vietnam, and to investigate antibiofilm activity of this compound against three Staphylococcus aureus (S. aureus) strains, one of which was methicillin-resistant S. aureus (MRSA) and the other two strains were methicillin-sensitive S. aureus (MSSA).
AMG in n-hexane fraction was isolated on a silica gel column and chemically analyzed by HPLC and NMR. The antibiofilm activity of this compound was investigated by using a 96-well plate model for the formation of biofilms. Biofilm biomass was quantified using crystal violet. The viability of cells was observed under confocal microscopy using LIVE/DEAD BacLight stains. Biofilm composition was determined using specific chemical and enzyme tests for polysaccharide, protein and DNA. Membrane-damaging activity was assayed by measuring the hemolysis of human red blood cells in presence of AMG.
The results indicated that the isolated AMG, with a purity that exceeded 98%, had minimal inhibitory concentrations in the range of 4.6-9.2 μmol/L for the three strains tested. Interestingly, the MSSA strains were more sensitive to AMG than the MRSA strain. Minimal bactericidal concentrations were 2-fold higher than the minimal inhibitory concentration values for the three strains, indicating that AMG was a bactericidal compound. AMG also prevented biofilm formation effectively, albeit that again the MRSA strain was the most resistant. Interestingly, biofilms of the MRSA strain contained protein as a main component of the extracellular matrix, whereas this was polysaccharide in the MSSA strains. This might relate to the resistance of the MRSA 252 strain to AMG. Assays using human red blood cells indicated that AMG caused significant membrane damage with 50% of cell lysis occurred at concentration of about 36 μmol/L.
Our results provide evidence that the isolated AMG has inhibitory activity against biofilm formation by S. aureus, including MRSA. Thus, isolated AMG proposes a high potential to develop a novel phytopharmaceutical for the treatment of MRSA.
从越南种植的山竹果皮中分离出α-倒捻子素（AMG），并研究该化合物对三株金黄色葡萄球菌的抗生物膜活性，其中一株为耐甲氧西林金黄色葡萄球菌（MRSA），另外两株为甲氧西林敏感金黄色葡萄球菌（MSSA）。
通过硅胶柱分离正己烷馏分中的AMG，并采用高效液相色谱法（HPLC）和核磁共振波谱法（NMR）进行化学分析。利用96孔板模型研究该化合物的抗生物膜活性。使用结晶紫对生物膜生物量进行定量。在共聚焦显微镜下使用LIVE/DEAD BacLight染色观察细胞活力。通过对多糖、蛋白质和DNA进行特定的化学和酶测试来确定生物膜的组成。通过测量AMG存在下人红细胞的溶血情况来测定膜损伤活性。
结果表明，分离得到的AMG纯度超过98%，对三株受试菌株的最低抑菌浓度在4.6 - 9.2 μmol/L范围内。有趣的是，MSSA菌株比MRSA菌株对AMG更敏感。最低杀菌浓度比三株菌株的最低抑菌浓度值高2倍，表明AMG是一种杀菌化合物。AMG也能有效防止生物膜形成，不过MRSA菌株的抗性最强。有趣的是，MRSA菌株的生物膜含有蛋白质作为细胞外基质的主要成分，而MSSA菌株的生物膜主要成分是多糖。这可能与MRSA 252菌株对AMG的抗性有关。用人红细胞进行的试验表明，AMG会导致显著的膜损伤，在浓度约为36 μmol/L时发生50%的细胞裂解。
我们的结果提供了证据，表明分离得到的AMG对包括MRSA在内的金黄色葡萄球菌生物膜形成具有抑制活性。因此，分离得到的AMG具有开发用于治疗MRSA的新型植物药的巨大潜力。
