Использование субкритической воды в качестве подвижной фазы при анализе методом ВЭЖХ

Abstract

 The aim of the study was an investigation into the possibility of using hot and subcritical water as a mobile phase in HPLC analysis of a mixture of aromatic substances, as well as milk thistle extract components, on columns with hypercrosslinked polystyrene MMN1 and silica gel modified with octadecyl groups.

The HPLC analysis of a model mixture of benzene and toluene 1:1 by volume on a Hypersil GOLD C18 column showed similar elution strength for a mixture of acetonitrile : water 20:80 (% vol.) and hot water at a temperature of 90°C. Water at a temperature of 125°C according to the elution characteristics corresponded to the ratio of acetonitrile: water 30:70 (% vol.). The number of theoretical plates when changing the water-acetonitrile mobile phase to subcritical water indicates a decrease in its efficiency by about 25-30%. The same trend was observed in the HPLC analysis of other analytes using subcritical water as the mobile phase. Due to the formation of the fluid structure of water under subcritical conditions, the pressure in the system changed, which allows varying the flow rate of the eluent, which partly compensates for the smearing of the peaks. An aqueous extract of milk thistle, obtained under subcritical conditions at a temperature of 170°C, was chosen as an object for studying the elution characteristics of hot water on a Hypersil GOLD C18 column. The experiment was carried out at temperatures of 90 and 125°C. The analysis temperature was not further increased due to the deterioration of the separation of the analyte components, and also because of the low thermal stability of the C18 stationary phase. Optimum results under the experimental conditions were obtained at a temperature of 90°C, while complete separation could not be achieved.

The model mixture of benzene: toluene: xylene was not eluted with a mobile phase of acetonitrile: water 20:80 (vol.%) by HPLC analysis on a Purolite MMN1 column under standard conditions. With an increase in the concentration of acetonitrile in the mobile phase to 80%, the elution of the components also did not occur. The use of subcritical water as a mobile phase at temperatures of 125, 150, 170°C did not lead to elution of the components from the Purolite MMN1 column. The milk thistle extract was separated on a Purolite column with a MMN1 sorbent under standard conditions, while optimal results were obtained with the mobile phase acetonitrile: 0.01M phosphate buffer (pH=3) at a ratio of 35:65. The components of the milk thistle extract - silybin, silicristin, silydianin, and taxifolin are poorly soluble in water, have benzene rings in their structure, but the size and polarity of the molecules allow to avoid irreversible interactions with the stationary phase under the conditions of analysis, as in the case of a model mixture, possibly in this case a molecular sieve effect is observed.