| Plant hormones play a very important role in the regulation of plant development, they can be divided into the following five categories: auxin(IAA), cytokinin(CKS), abscisic acid(ABA), gibberellin(GAs) and ethylene(ET) according to their structure and physiological function. They play a key role in the life cycle of plants, including cell division, expansion and differentiation, organ development, seed dormancy and germination as well as leaf senescence and abscission. Plant hormones in plant tissues are ultra trace level, usually in pg/g- ng/g fresh weight(FW) range, and around the coexistence of very complex matrix composition at the same time, it is impossible to analyze all of plant hormones. Therefore,it has become one of the important problems in the study of the mechanism of the action of plant hormones to make qualitative and quantitative analysis of the plant hormones.Chromatographic techniques have been widely used for its simple, rapid and highly efficient features.Chromatography coupled with mass spectrometry, in particular, liquid chromatography and mass spectrometry methods not only provide the structural information based on fragmentation patterns, but also significantly improve the sensitivity. based on the above theory, the method of high performance liquid chromatography and high performance liquid chromatography coupled with mass spectrometry for the detection of plant hormones was established. The content of plant hormones and organic acid in vegetables and fruits have been detected, and the results are satisfied:1. Analysis of IAA, IBA, NAA and ABA by HPLCA high performance liquid chromatography(HPLC) method was established for the separation and determination of indole-3-acetic acid(IAA), indole butyric acid(IBA), abscisic acid(ABA), andnaphthalene acetic acid(NAA). Using Eclipse XDB-C8(4.6 mm*250 mm, 5 ?m)reversed-phase column as seperation column. The mobile phase composed of 70% methanol and 30% buffer(formic acid and ammonium formate solution, pH=3.2)=70:30, the flow rate was 1 mL/min, the injection volume was 5 ?L,the temperature of the column is 25 ?, The detector wavelength is 280 nm,The results showed that the separation effect of the four hormones was satisfactory, and the correlation coefficient was in the linear range for 0.9979-0.9983, The detection limit was 0.03-0.1?g/m L, the recovery rate reached95.8 %-104.6 % of standard addition, The relative standard deviation was not higher than 0.53%. The developed method was appied to analysis real samples, 3- indole acetic acid in bean sprouts and abscisic acid in pear were 16.8 mg/kg and 5.0 mg/kg.2.Analysis of GA3 and GA12 by HPLC-MS methodThis method uses Hilic C18(2.1 mm*50 mm, 2.6 ?m) column, V(methanol):V(0.01 mol/L ammonium acetate) =80:20, separation of flow rate at 0.2 mL/min,by using electrospray ionizationinmultiple-reactionmonitoringmode, In order to obtain higher sensitivity, a multiple reaction monitoring mode was used. The parent ions of GA3 and GA12 were m/z 345.3 and 331.4, and the daughter ions were m/z at 143, 220.9 and 201, 269.2, respectively. The results showed that the establised method has a good linear relationship(R2>0.99)in the 0.05-10 ?g/mL linear range, the detection limit was 0.01 ?g/mL,andthe average recovery rate was between 81.7 % and 103.2 % in three supplemental level of 0.20, 1.5, 8.0?g/mL withrelative standard deviation(RSD) was between 1.9 % and 0.26 %.3. Analysis of organic acid by LC-MS method with chemical derivatizationIn this section, a LC-MS method for the analysis of malic acid and citric acid with chemical derivatization was developed. The weigh of organic acids, Na2CO3,2- bromoacetophenone(molar ratio1:1:1) 500 ?L to 50 mL flask,mix evenly, reaction of 24 h under 55 ? oil bath, qualitative analysis of reactants after cooling of reaction, Using reversed phase column Agilent C18(2.1 mm*150 mm, 5 ?m), V(methanol):V(0.1% acetic acid aqueous solution) = 60:40 as mobile pahse. The flow rate was 0.2 mL/min,the injection volume was 5 ?L,by using electrospray ionizationin positive ion monitoringmode. The results showed that the correlation coefficients of the two kinds of organic acids in the linear range after derivatization was 0.9754-0.9966,detection limit for each was 6.67 ?g/ml, 2.33 ?g/ml, the recovery rate was between 81.9%-91.0%. The malic acid in apple and citric acid in lemon were measured by the derivative method of 15.3 ?g/mL 19.4 ?g/mL. |
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