The influences of the organic component of the mobile phase and the column temperature on the retention of ginsenosides on a poly(vinyl alcohol) (PVA) bonded stationary phase operated under hydrophilic interaction chromatographic mode were investigated. The retention of the ginsenosides was found to increase with increasing amount of acetonitrile (MeCN) in the mobile phase, which is typical of hydrophilic interaction chromatographic behavior. It was also found that the retention of the analytes was highly affected by the type of the organic modifier used. Aqueous MeCN (75-90%) gave the most satisfactory retention and separation of ginsenosides Rf, Rg1, Rd, Re, Rc, Rb2 and Rb1 compared with aqueous methanol, isopropyl alcohol or tetrahydrofuran at the same composition levels. The effects of the different types of organic modifiers on the retention of the analytes were attributed to their solvent strength and hydrogen-bond accepting/donating properties. The effect of temperature on the retention of ginsenoside on the PVA-bonded phase was assessed by constructing van't Hoff plots for two temperature ranges: subambient (273-293 K) and ambient-elevated (298-333 K) temperatures. van't Hoff plots for all analytes were linear at the two temperature intervals; however, the slopes of the lines corresponding to ginsenosides Rg1 and Re were completely different from those for the rest of the analytes especially in the subambient temperature range. Enthalpy-entropy compensation (EEC) studies were conducted to verify the difference in thermodynamics observed for ginsenosides Rg1 and Re compared with the other analytes. EEC plots showed that Rf, Rd, Rc, Rb2 and Rb1 were possibly retained by the same retention mechanism, which was completely different from that of Rg1 and Re at subambient temperatures. Retention prediction models were derived using multiple linear regression to identify solute attributes that affected the retention of the analytes on the PVA-bonded phase. The mathematical models derived revealed that the number of hydrogen-bond donors and the ovality of the molecules are important molecular properties that govern the retention of the compounds on the chromatographic system.
The chemical composition of bufadienolides isolated from the venom of Bufo viridis green toad occurring in Central Asia was determined and their biological properties were studied. Six individual bufadienolides were isolated by reverse-phase chromatography on a Lichrosorb RP-8 (10 m) column in amounts sufficient for qualitative analysis. Two of these were previously identified as arenobufagin and gamabufotalin by NMR spectroscopy and x-ray diffraction methods. The chemical structures of four other bufadienolides are now established by NMR spectroscopy and HPLC. These compounds have been identified as telocinobufagin (3b,5b,14b-trihydroxybufa-20,22-dienolide), marinobufagin (3b,5b-hydroxy-14,15b-epoxybufa-20,22-dienolide), bufarenogin (3b,12b,14b-hydroxybufa-20,22-dienolide), and bufalin (3b,14b-hydroxybufa-20,22-dienolide).
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