Development of Gradient Centrifugal Partition Chromatography Method and Its Application for the Isolation of 3,5-Dimethoxyphenanthrene-2,7-diol and Batatasin-I from Dioscorea opposita원문보기
Gradient centrifugal partition chromatography (GCPC) method was developed and applied to isolate 3,5-dimethoxyphenanthrene-2,7-diol (DMP) and batatasin-I (BA-I) from the dichloromethane soluble extract of Dioscorea opposita. In this method, the lower phase of n-hexane-methanol-water system (HMW, 10 ...
Gradient centrifugal partition chromatography (GCPC) method was developed and applied to isolate 3,5-dimethoxyphenanthrene-2,7-diol (DMP) and batatasin-I (BA-I) from the dichloromethane soluble extract of Dioscorea opposita. In this method, the lower phase of n-hexane-methanol-water system (HMW, 10 : 9 : 1, v/v) was used as a mobile phase A (MpA) and water was used as a mobile phase B (MpB). This gradient CPC method is comparable to that of reversed-phase HPLC method in that the stationary upper-phase of HMW (10 : 9 : 1 v/v) works as if it were reversed-phase silica gel due to its hydrophobic property, while the lower phase (MpA) and water (MpB) functioned as hydrophilic mobile phases. The initial condition of the mobile phase was 20% MpA/80% MpB and maintained for 150 min to obtain DMP (1.2 mg), and then MpA was increased up to 50% to elute BA-I (1.7 mg). The purities of DMP and BA-I were 94.1% and 98.3% with the recovery yields of 83% and 86%, respectively. Similar results were obtained by linear-gradient CPC. The CPC peak fractions were identified by comparing their retention time to those of authentic samples of DMP and BA-I and their spectroscopic data ($^1$H NMR and $^{13}$C NMR) to those of literature values.
Gradient centrifugal partition chromatography (GCPC) method was developed and applied to isolate 3,5-dimethoxyphenanthrene-2,7-diol (DMP) and batatasin-I (BA-I) from the dichloromethane soluble extract of Dioscorea opposita. In this method, the lower phase of n-hexane-methanol-water system (HMW, 10 : 9 : 1, v/v) was used as a mobile phase A (MpA) and water was used as a mobile phase B (MpB). This gradient CPC method is comparable to that of reversed-phase HPLC method in that the stationary upper-phase of HMW (10 : 9 : 1 v/v) works as if it were reversed-phase silica gel due to its hydrophobic property, while the lower phase (MpA) and water (MpB) functioned as hydrophilic mobile phases. The initial condition of the mobile phase was 20% MpA/80% MpB and maintained for 150 min to obtain DMP (1.2 mg), and then MpA was increased up to 50% to elute BA-I (1.7 mg). The purities of DMP and BA-I were 94.1% and 98.3% with the recovery yields of 83% and 86%, respectively. Similar results were obtained by linear-gradient CPC. The CPC peak fractions were identified by comparing their retention time to those of authentic samples of DMP and BA-I and their spectroscopic data ($^1$H NMR and $^{13}$C NMR) to those of literature values.
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opposita, and then MpA was increased up to 50% to get BA-I. As a result, DMP (peak fraction 1, 45 - 110 min, 1.2 mg) and BA-I (peak fraction 2, 290 - 430min, 1.7 mg) were successfully isolated, and the purities of DMP and BA-I were 94.1% and 98.3%, respectively (Fig. 5). At a glance, the absolute amounts of isolated compounds seemed to be very small because the content levels of DMP and BA-I are very low in dichloromethane soluble extract of D.
5). At a glance, the absolute amounts of isolated compounds seemed to be very small because the content levels of DMP and BA-I are very low in dichloromethane soluble extract of D. opposita, but the recovery yields of DMP and BA-I were 83% and 86%, respectively, determined by quantitative HPLC analysis. Similar results were obtained in linear-gradient CPC (Fig.
opposita, but the recovery yields of DMP and BA-I were 83% and 86%, respectively, determined by quantitative HPLC analysis. Similar results were obtained in linear-gradient CPC (Fig. 4B) and these results reveal that the upper- and lower phase of HMW (10 : 9 : 1, v/v) and water are very effective to isolate and purify DMP and BA-I from dichloromethane soluble extract of D. opposita with high purities and recovery yields. Furthermore, this solvent system can be utilized to fractionate or isolate diverse natural products by altering the polarity of mobile phase like reversed-phase column chromatography.
opposita. The recovery yields of DMP and BA-I were 83% and 86%, respectively. The results of the present study clearly demonstrate that CPC with gradient elution can provide reference compounds from D.
The recovery yields of DMP and BA-I were 83% and 86%, respectively. The results of the present study clearly demonstrate that CPC with gradient elution can provide reference compounds from D. opposita, and the two-phase solvent system composed of HMW (10 : 9 : 1, v/v) and water can be utilized as an alternative method for the fractionation or isolation of diverse natural products instead of solid supported column chromatography.
The retention of stationary phase remained satisfactory level at 68.4 ± 2.1% (0 - 100% of MpA), and the variation was less than 9.7% indicating that the solvents composed of the upper- and lower-phase of HMW (10 : 9 : 1, v/v) and water are very stable system for gradient CPC (Fig. 3).
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