肠道菌群模式菌株的HPLC分离条件优化及其表征
Optimization of separation conditions and characterization of the model strains of intestinal microflora by high performance liquid chromatography

采用高效液相色谱分别考察了肠道菌群模式菌株及各单菌的色谱行为，成功建立了HPLC分离分析肠道菌群模式菌株的最佳洗脱方法，即阶梯式梯度洗脱. 结果显示，肠道模式混合菌株经过高效液相色谱分离得到5个明显的洗脱组分，分别为0.10mol·L-1（脆弱拟杆菌和金黄色葡萄球菌）、0.20mol·L-1（青春双歧杆菌和鼠李糖乳杆菌）、0.25mol·L-1（粪肠球菌）、0.30mol.L-1（奇异变形杆菌、克雷伯肺炎杆菌和产气荚膜梭菌）和0.50mol·L-1（大肠杆菌和产气荚膜梭菌）洗脱峰. 色谱柱的上样浓度为培养菌液（OD600=0.576）的5倍时，其最大上样量为500μL，且方法分辨率高、重现性好、结果稳定.
High performance liquid chromatography (HPLC) was used to investigate the chromatographic behavior of the mixture of 9 typical intestinal microflora and single strain therein，respectively，and established a step gradient elution method of HPLC for separation and analysis of the mixed strains. The results showed that the mixture of 9 typical intestinal microflora was efficiently separated by HPLC and 5 eluting fractions were obtained，which contained 0.10mol·L-1 (B. fragilis and S.aureus)，0.20mol·L-1 (B. adolescentis and L. rhamnosus)，0.25mol·L-1 (E. faecalis)，0.30mol·L-1 (P. mirabilis，K. Pneumoniae and C. perfringens)，and 0.50mol·L-1 (E. coli and C. perfringens) fractions. The maximum load of sample was 500μL of 5-fold concentrate of the bacterial broth with an OD600 of 0.576，and the HPLC method presented here has high resolution，good reproducibility and stable results