Determination of 8 Glycosides and 4 Aglycones in Directionally Processed Astragalus membranaceus under Different Temperatures by UPLC-MS / 中国药房

ABSTRACT

OBJECTIVE：To establish a method for the determination of 8 glycosides（astragaloside Ⅰ，Ⅱ，Ⅲ，Ⅳ and calycosin glucopyranoside ，2′-hydroxy-3′，4′-dimethoxy-isoflavan-glucoside and 9，10-dimethoxy-pterocarpan-glucoside） and 4 aglycones（calycosin，formononetin，7，2′-dihydroxy-3′，4′-dimethoxy-isoflavan and 3-hydroxy-9，10-dimethoxy-pterocarpan） in Astragalus membranaceus ，and to investigate the effects of different processing temperatures on the contents of above 12 components. METHODS ：The contents of 12 components in A. membranaceus and samples processed under different temperatures（120，140，160，180，200 ℃）were determined by UPLC-MS/MS. The determination was performed on ACQUITY UPLC HSS T 3 column with mobile phase consisted of 0.1 mol/L formic acid water solution -0.1 mol/L formic acid acetonitrile solution （gradient elution ）at the flow rate of 0.5 mL/min. The column temperature was 30 ℃. The detection wavel ength was 260 nm，and sample size was 2 μL. Electrospray ion source（ESI）was used under positive ion mode （ESI+）. The mass scanning range was mass ratio （m/z）of 50-1 500，with capillary voltage of 2 000 V and ion source temperature of 100 ℃. The desolvation temperature was 400 ℃；flow rate of atomizing gas （N2） was 40 L/h，and that of desolvation was 800 L/h；collision energy （CE）was 20-30 V；data acquisition rate was 0.5 s/scan. RESULTS：The linear range of astragaloside Ⅰ，astragaloside Ⅱ，astragaloside Ⅲ，astragaloside Ⅳ，calycosin-glucopyranoside， calycosin，ononin，formononetin，2′-hydroxy-3′，4′-dimethoxy-isoflavan-glucoside，7，2′-dihydroxy-3′，4′-dimethoxy-isoflavan，9， 10-dimethoxy-pterocarpan-glucoside and 3-hydroxy-9，10-dimethoxy-pterocarpan were 0.001 16-0.232 0，0.000 276-0.055 2， 0.000 22-0.044 0，0.000 225-0.045 0，0.000 734-0.587 0，0.001 17-0.234 0，0.000 742- 0.148 0，0.001 30-0.260，0.003 98-0.795 0， 0.000 476-0.476 0，0.001 89-0.378 0，0.000 336-0.336 0 μg（all R2≥0.999 2），respectively. The limits of detection were 6.2×10－6， 4.8×10－6，3.8×10－6，3.4×10－6，5.8×10－6，4.8×10－6，4.2×10－6，3.2×10－6，5.8×10－6，2.6×10－6，4.2×10－6，6.4×10－6 μg，respectively. The limits of quantitation were 12.6×10－6，16.2×10－6，14.4×10－6，14.8×10－6，18.8×10－6，16.4×10－6，15.4×10－6，10.8×10－6，20.2×10－6， 12.4×10－6，14.6×10－6，23.4×10－6 μg，respectively. RSDs of precision ，stability（24 h）and repetition tests were all lower than 3.0％（n＝6）. The average recoveries were 99.1％，100.2％，98.7％，101.9％，98.6％，102.1％，99.2％，100.3％，98.7％， 99.2％，99.3％ and 100.8％，with the RSDs of 1.9％，2.2％，2.4％，1.8％，2.1％，1.7％，2.3％，1.9％，2.4％，1.8％，2.2％ and 1.9％（n＝6），respectively. The results showed that the contents of astragaloside Ⅰ，Ⅱ and Ⅲ decreased gradually with the increase of processing temperature ；the content of astragaloside Ⅳ increased gradually with the increase of temperature. The content of flavonoid glycosides ，such as calycosin glucopyranoside ，ononin，2′-hydroxy-3′，4′-dimethoxy-isoflavan-glucoside and 9， 10-dimethoxy-pterocarpan-glucoside decreased with the increase of temperature ；the corresponding aglycone components as flavonoid glycosides ，formononetin，3-hydroxy-9，10-dimethoxy- pterocarpan increased firstly and then decreased with the increase ； the content of 7，2′-dihydroxy-3′，4′- dimethoxy-isoflavan decreased with the increase of temperature. CONCLUSIONS ：Established UPLC-MS/MS method can be used for determination of 12 components in A. membranaceus . After processed under different temperature，the contents of glycosides decreased in general ，while the contents of aglycones increased in general.