The Study Of Synthesis And Application Of New Type Of Carboxyl Bonded Silica Gel Stationary Phase For HPLC

Since the establishment of HPLC, the study of chromatographic stationary phase has become an important area for chromatography researchers. In the past few years, because of people's renewed attention on the Chinese herbal medicine, traditional Chinese herbs have got a rapid development and widely used in prevention and treatment of a variety of diseases. But the structures of traditional Chinese herbs are complicated, it demands some chromatography stationary phases which have kinds of functions separating them. As the active ingredients of Chinese Herbs have their own unique structure, it can play a role with a variety of substances, which could be used as separation materials of Chinese herbal medicine, a series of new high performance liquid chromatography bonded stationary on Chinese herbal ingredients were prepared. On the basis of the previous literatures, we prepared two kinds of bonded stationary phases which all used rhein as the raw material in this dissertation. Their chemical structures were characterized, and the chromatographic performances of them were also investigated. The chromatographic separation mechanism was initially discussed, and some chromatographic exploratory developments were done on the two new stationary phases. The major content contains:1.γ-aminopropyltriethoxy silane was used as the coupling agent to prepare a amide rhein-bonded silica gel stationary phase.2. y-glycidoxypropyltrimethoxy silane was used as the coupling agent to prepare a ester rhein-bonded silica gel stationary phase.3. The new amide rhein-bonded silica gel stationary phase was used to separate two isoflavone compounds under the simple mobile phase of methanol-water.4. The separation performance of the new ester rhein-bonded silica gel stationary phase was researched by two bio-basic groups.1. Simple preparation and evaluations of amide rhein-bonded silica gel stationary phaseA new amide carboxy bonded silica gel stationary phase (RBSP) was prepared which was used rhein as the raw material and y-aminopropyltriethoxy silane as the coupling agent. The structure of RBSP was characterized by infrared spectroscopy, thermal analysis and elemental analysis. The effect of methanol content in the mobile phase on chromatographic performance of the bonded stationary phase was investigated. The chromatographic performance of RBSP was also evaluated using the mixture of acidic, neutral and basic compounds as solute. The separating property of bonded stationary phase was carried out using methanol-water as the mobile phase by separating two kinds of soy isoflavones and several biological bases compared with C18. The chromatographic separation mechanism was also initially discussed. The results demonstrated that RBSP possessed good reversed-phase chromatographic behavior. At the same time, because of the bonded stationary phase containing phenolic hydroxyl and amide groups which can provide action sites for multiple compounds, it has the obvious advantage in separating polar compounds especially the fast separating rate. So the RBSP could be efficiently used for the separation of polar compounds.2. Preparation and evaluations of ester rhein-bonded silica gel stationary phaseIn this study, y-glycidoxypropyltrimethoxy silane was used as the coupling agent reacting with rhein to prepare a ester rhein-bonded silica gel stationary phase(RESP). The structure of RESP was characterized by multiple methods. And the chromatographic performance was also evaluated using various solutes compared with C18. The results indicated that the bonded stationary phase can hold-back the alcohol groups of silica gel because of the esterfunction and anthraquinones on RESP. So it could separate many compounds efficiently.3. The analytical application of amide rhein-bonded silica gel stationary phase to two isoflavone compounds.A amide rhein bonded silica gel stationary phase (RBSP) was used for the separation of two isoflavones which named daidzin and glycitin, and the chromatographic behavior of the two isoflavones was studied in comparison with conventional C18. The retention behavior of the two isoflavones on this stationary phase was investigated through changing the construction of mobile phase.The experiments showed that under the same chromatographic condition, RBSP could tacho-separate the two isoflavones when using methanol-water (V/V=75/25) as the mobile phase, but the elution order opposited compared to C18. And the results illustrate that RBSP is different from C18 at the retention mechanism.4. Separation and determination of two bio-basic groups on the ester rhein-bonded silica gel stationary phase.The analytical research was carried out by comparing the chromatographic behavior of uracil and adenine on the new bonded stationary phase (RESP) with the conventional C18.It could efficiently separate the two basic groups under the optimized chromatographic conditions. And uracil and adenine in urine were determined on RESP. It was found that the elution order of the two basic groups on RESP was unanimously compared to C18. It indicates that the separation mechanism of RBSP is mainly in view of hydrophobic interaction.