反相高效液相色谱柱前衍生法测定发酵液中的5-氨基乙酰丙酸  

Determination of 5-Aminolevulinic Acid in E.coli Fermentation Broth by Reversed Phase High Performance Liquid Chromatography with Precolumn Derivatization

作　　者：范理[1] 林建平[1] 刘晓侠[1] 秦刚[1] 岑沛霖[1] 

机构地区：[1]浙江大学生物工程研究所,杭州310027

出　　处：《分析化学》2006年第7期937-940,共4页Chinese Journal of Analytical Chemistry

基　　金：国家自然科学基金(No.20306026);重大基础研究前期研究专项(No.2004CCA5500)资助

摘　　要：建立了同时检测发酵液中5-氨基乙酰丙酸和甘氨酸的柱前衍生反相高效液相色谱法.使用ZORBAX SB-AQ型色谱柱（250 mm×4.6 mm i.d.,5 μm）,以甲醇-50 mmol/L醋酸钠缓冲溶液（pH 6.2）作为流动相梯度洗脱.流速为1 mL/min,柱温25℃,254 nm波长下检测30 min.在此条件下,5-氨基乙酰丙酸和甘氨酸在浓度为0.8～23 mmol/L和1.3～66 mmol/L时与色谱峰面积值之间线性关系良好,回归系数均在0.9997以上.发酵液产物的加标回收率在94.9%～104.7%.将检测结果与分光光度法相比,符合较好.测试数据表明,此方法简单、可靠,对于发酵产物的分离效果好.A reversed phase high performance liquid chromatography with precolumn derivatization was developed for the simultaneous determination of 5-aminolevulinic acid （ALA） and glycine （Gly） in E. coli fermentation broth. A ZORBAX SB-AQ column （250 mm ×4.6 mmi. d. ,5μm） was used for separation, and the elution was performed with linear gradient elution of methanol and 50 mmol/L sodium acetate solution （ adjustment of pH to 6.2 with acetic acid ）. The flow rate was 1 mL/min, the temperature of column was 25 ℃, and the detection wavelength was at 254 nm with the detection time of 30 minutes. The dynamic linear range is 0.8 - 23 mmol/L and 1.3 - 66 mmol/L for ALA and Gly respectively, and the correlation coefficients are both above 0.9997. The average recoveries of ALA in 3 samples of fermentation broth ranged from 94.9% to 104. 7%. The results agreed well with those of spectrophotometry. So the developed method is simple and reliable.

关 键 词：5-氨基乙酰丙酸 反相高效液相色谱法 梯度洗脱 柱前衍生 发酵液 

分 类 号：O657.72[理学—分析化学] TS207.3[理学—化学]