杨树嫩茎质外体中脯氨酸含量测定方法的建立

采用高效液相色谱法HPLC对微透析液进行痕量脯氨酸的检测,研究了痕量脯氨酸柱前衍生化的方法。采用苯异硫氰酸酯PITC-乙腈溶液和三乙胺-乙腈溶液对透析液中痕量脯氨酸进行衍生化后,通过环己烷萃取,得到透析液中痕量脯氨酸衍生化的最适方法,无需后续处理可直接进样,并确定了HPLC检测条件:岛津Inertsil ODS-SP色谱柱(250 mm×4.6 mm, 5 μm);柱温:31 ℃;流动相A:0.05 mol·L^-1醋酸钠溶液;流动相B:乙腈-流动相A(1:1);洗脱条件:A:B(50:50);流速:1.0 mL·min^-1;检测波长:254 nm;进样量:20 μL。脯氨酸加标回收率为95.1 ％~104.2 %。同时,对3种杨树质外体透析液中的脯氨酸进行了测量。

Determination of Proline Content in Apoplast of Poplar Stems

Based on microdialysis experience in apoplast of poplar stems, the trace of proline in plant was detected by using the High-performance liquid chromatography (HPLC) method, precolumn derivatization method was studied. Phenyl isothiocyanate PITC- acetonitrile solution and triethylamine acetonitrile solution were applied to derivate the proline. Cyclohexane was used to extract proline, the sample could be directly injected in HPLC without subsequent processing. The detection conditions were determined: the SHIMADZU Inertsil ODS-SP column (250 mm×4.6 mm, 5 μm); column temperature: 31 ℃; mobile phase A: 0.05mol·L^-1 sodium acetate solution, mobile phase B: acetonitrile-mobile phase A (1:1); Elution conditions: A:B (50:50); flow rate: 1.0 mL·min^-1; detection wavelength: 254nm; the injection volume was 20μL. The recovery rate of proline was 95.1-104.2%. At last the proline contents of three poplar apoplast dialysis were detected by this method