超临界流体色谱法分析非对映异构体d4T-5’-N-磷酰化苯丙氨酸甲酯

摘要

采用超临界CO2流体色谱技术,分析d4T-5'-N-磷酰化苯丙氨酸甲酯手性磷的非对映异构体.色谱柱为Hpersil ODS2 (250 mm× 4.6 mm,5 μm),流动相为夹带改性剂甲醇、乙醇和异丙醇的超临界CO2流体.以容量因子、选择性和分离度为指标,考察改性剂、背压和柱温对分离的影响.在甲醇、乙醇和异丙醇3种改性剂中,甲醇为最好的改性剂,其中在7％甲醇改性剂下,该化合物的分离度可达到3.35.在7％甲醇改性剂条件下,考察了压力( 10～20 MPa)和温度(303.15～318.15 K)的影响.在优化的分离条件(改性剂为7％甲醇,流速为2 mL/min,柱温为308.15 K,背压为15 M Pa)下,d4T-5'-N-磷酰化苯丙氨酸甲酯的两种非对映异构体完全达到基线分离,分离时间约15 min.%The separation of phenylalanine methylester 5'-phosphoamidates of d4T ( d4T-P-N-PheOMe) diastereoisomers has been investigated using supercritical fluid chromatography on Hypersil ODS2 (250 mm×4. 6 mm, 5 μm). Supercritical CO2 modified with methanol, ethanol and 2-propanol was used as mobile phase. The influence of type and concentration of alcohol modifier, back pressure and column temperature on retention factor, diastereoselectivity and resolution was studied. Among methanol, ethanol and 2-propanol, methanol was proved to be the most favorable modifier, and 7% (V/V) of methanol was the preferred concentration at which d4T-P-N-PheOMe diastereoisomers could be separated with resolution of 3. 35. The effects of pressure and temperature were investigated at 7% (V/V) of methanol in the pressure range of 10 - 20 MPa and temperature range from 303. 15 to 318. 15 K. The optimal separation condition was 7% modifier in methanol as mobile phase, 2. 0 mL/min of flow rate for mobile phase, 308. 15 K of column temperature, 15 MPa of back pressure. d4T-P-N-PheOMe could be separated baselinely and the running time was about 15 min under this optimal separation condition.