RESUMO
In the present study, the long-term high-temperature (>80 degrees C) and temperature programming stability of fused silica capillaries packed with 5 microm PLRP-S 300 A and monolithic PS-DVB capillaries (both 180 microm id x 6 cm) under reversed-phase conditions has been examined. In isothermal mode, the columns were defined as temperature-stable when a less than 10% change in apparent retention factors (k) and a less than 20% change in "retention time/peak width"-factors (n) of the probe solutes (proteins) were observed after passing 7,500 void volumes of effluent through the columns (about 100 h operation). According to these criteria, the PLRP-S and monolithic capillaries were defined temperature-stable at 100 and 130 degrees C, respectively. Furthermore, when continuously running temperature programs between 50 degrees C and the upper temperature limit determined in isothermal mode, virtually no change in k or n were observed on neither of the columns after running more than 35,000 void volumes or 1,600 temperature programs. Additionally, temperature-programmed reversed-phase separations of proteins on both types of capillaries are demonstrated and discussed.
RESUMO
In the present study, three different methods for packing of 3 microm PBD-ZrO2 particles in 0.5 mm i.d. glass-lined stainless steel columns have been examined. The two first methods were based on a traditional downstream high-pressure technique using tetrachloromethane (Method I) or aqueous Triton X-100 (Method II) as slurry solvents, while Method III was an upstream high-pressure flocculating method with stirring, using isopropanol both as the slurry and packing solvent. Method I was found to be superior in terms of efficiency, producing 0.5 mm i.d. x 10 cm columns with almost 90,000 plates m(-1) for toluene (R.S.D. = 8.7%, n = 3), using a slurry concentration of 600 mg ml(-1), ACN-water (50:50 (v/v)) as the packing solvent and a packing pressure of 650 bars. For Method I, the slurry concentration, column i.d., column length and initial packing pressure were found to have a significant effect on column efficiency. Finally, the long-term temperature stability of the prepared columns was investigated. In isothermal mode, using ACN-20 mM phosphate buffer, pH 7 (50:50 (v/v)) as the mobile phase, the columns were found to be stable for at least 3,000 void volumes at 100 degrees C. At this temperature, the solute efficiencies changed about 5-18% and the retention factors changed about 6-8%. In temperature programming mode (not exceeding 100 degrees C), on the other hand, a rapid decrease in both column efficiency and retention factors was observed. However, when the columns were packed as initially described, ramped up and down from 50 to 100 degrees C for 48 h and refilled, fairly stable columns with acceptable efficiencies were obtained. Although not fully regaining their initial efficiency after refilling, the solute efficiencies changed about 19-28% (32-37%) and the retention factors changed about 4-5% (13-17%) after running 3,000 (25,000) void volumes or 500 (3,900) temperature programs.
