The Analysis Method Of Soyasaponins And Its Diversity Analysis

Soyasponins is the secondary metabolites of beans, which have varieties of benefits for human body and have multiple defence functions in plants. This paper established a method to detect soyasponins rapidly use TLC, a method to detect soyasponins use HPLC-ESI-MS/MS and a method to detect soyasponins use HPLC.The condition of TLC was chloroform:methanol:water:formicacid (65:38:10:0.01) which was placed a night for 30min and the TLC plates were developed in it for 70 minutes. The embryo and cotyledon of 20 kinds of soybean material and the seed coat, the root, steam, leaf were qualitative analysized using this method, the results showed that different materials have significant difference in band of TLC.The HPLC-ESI-MS/MS was established for detecting soybean saponin, the chromatographic conditions was that mobile phase A and B were 0.025% (v/v) acetonitrile solution of acetic acid and acetic acid aqueous solution, the temperature of column is 30?, the injection volume is 10 ?L, gradient elution time was 40 min, fog voltage is 3.5 kV, sheath and auxiliary gas pressure are 30 psi and 5 psi. According the standard of Aa, Ab and Ba, Bb of saponins to quantitate analysis. The results showed that the correlation coefficient of standard curve were greater than 0.997, the linear range is 10?1000 ?g/mL.Analyze the soyasaponin component and content of 46 kinds of seeds were from different ecological types in Shanxi used HPLC-ESI-MS/MS. It is indicated that exist significant difference for component and content of soyasaponin in different groups of the same material or between different materials. The average content of soyasaponins A in embryo, cotyledon and skin is 34.364,3.384,0.930 mg/g, the range of variation is 4.298?111.540 mg/g,0.020?9.881 mg/g,0?7.210 mg/g, the coefficient of variation is 81.08%,97.75%,181.61%, respectively. The average content of soyasaponins DDMP in embryo, cotyledon and skin is 60.631,3.272,0.149 mg/g, the range of variation is 1.545?50.507 mg/g,1.545? 8.056 mg/g,0.012?2.051 mg/g, the coefficient of variation is 108.45%,37.13%,242.95%, respectively. The average content of soyasaponins B in embryo, cotyledon and skin is 4.404,1.536,0.440 mg/g, the range of variation is 0.331?12.589 mg/g,0.008?1.925 mg/g,0.004?2.454 mg/g, the coefficient of variation is 87.24%,128.00%,136.59%, respectively.The average content of soyasaponins E in embryo, cotyledon and skin is 1.313,0.397,0.121 mg/g, the range of variation is 0.026?10.328 mg/g,0.026? 1.124 mg/g,0?1.239 mg/g, the coefficient of variation is 138.39%,62.97%,189.26%, respectively.The soybean samples were also classified into three groups on the basis of 100-seed weight that small (<13 g), medium (13-24 g), and large (>24 g), it is indicated that the content of soyasaponins B in embryo of small seeds is significantly higher than in large and medium seeds.The content of soyasaponin A in the cotyledons of small seeds is 1.82 and 1.87 times of the medium and large, DDMP is 1.51,1.64 times.The total content is 1.42,1.50 times.The soyasaponins of A, DDMP and DDMP+B+E in the small seeds is significantly higher than in large and medium seeds.The correlation analysis of soyasaponin found that soyasaponin in embryo exist highly significant positive correlation between A and the total amount of soyasaponin, the correlation coefficient is 88.8%, exist significant negative correlation between Aa and Ab. It is exist highly significant positive correlation between the total amount of soyasaponin and the total amount of soyasaponins A, DDMP and B, the correlation coefficient is 94.7%,50.3% and 63.7%, respectively. It is exist highly significant positive correlation between the total amount of soyasaponin DDMP and ag, ?g, yg in the embryo, the correlation coefficient is 90.8%,98.9% and 49.7%, respectively, it is exist highly significant positive correlation between the total amount of soyasaponin DDMP and ?g, Pa, ya in the cotyledon, the correlation coefficient is 54.5%,51.2% and 63.9%, respectively.The mobile phase of HPLC was consisted of (A) 0.05%(V/V) acetic acid in acetonitrile and (B) 0.05% acetic acid in water, and a timed gradient program was used.The solvent at a flow rate of 1.0 mL/min with detection at 205 nm for 100 min. The column temperature was at 30?, the injection volume is 30 ?L.The standards of soyasponins (Aa, Ab, Ba and Bb) are used as standard substance to establish standard carves,the linear range is 100?1000 ?g/mL. The results showed the coefficient of association of four standard carves are no less than 0.996, the average recovery rate is 89.9?105.7%, the precision is 1.06?2.31%. The detection limit of this method for soyasponins Aa, Ab is 20 ?g/mL, and Ba, Bb is 10 ?g/mL, but due to the lack of standards of soyasponins can't analyze the other saponins, the method still need to further improve.