Minimizing UV noise in supercritical fluid chromatography. I. Improving back pressure regulator pressure noise.

Pressure fluctuations and resulting refractive index changes, induced by the back pressure regulator (BPR) can be a significant source of UV detector noise in supercritical fluid chromatography (SFC). The refractive index (RI) of pure carbon dioxide (CO(2)) changes ≈0.2%/bar at the most commonly used conditions in supercritical fluid chromatography (SFC) (40 °C and 100 bar), compared to 0.0045%/bar for water (CO(2) IS 44× worse). Changes in RI cause changes in the focal length of the detector cell which results in changes in UV intensity entering the detector. The change in RI (ΔRI/bar) of CO(2) decreases 8-fold at 200 bar, compared to 100 bar. A new back pressure regulator (BPR) design representing an order of magnitude improvement in the state of the art is shown to produce peak to peak pressure noise (PN(p-p)) as low as 0.1 bar, at 200 bar, and 20Hz, compared to older equipment that attempted to maintain PN(p-p)<1bar, at <5Hz. With this lower PN(p-p), changes in baseline UV offsets could be measured as a function of very small changes in pressure. A pressure change of ±1 bar at 100 bar, common with some older BPR's, produced a UV baseline offset >0.5 mAU. A pressure change of ±0.5 bar representing the previous state-of-the-art, resulted in a UV offset of 0.3m AU. Baseline noise <0.05 is required to validate methods for trace analysis. The new BPR, with a PN(p-p) of 0.1 bar, demonstrated UV peak to peak noise (N(p-p))<0.02 mAU with a >0.03 min (10Hz) electronic filter under some conditions. This new low noise level makes it possible to validate SFC methods for the first time.

Berger TA ，Berger BK 《-》

Minimizing ultraviolet noise due to mis-matches between detector flow cell and post column mobile phase temperatures in supercritical fluid chromatography: effect of flow cell design.

A mis-match between the post-column mobile phase temperature and the UV detector flow cell temperature can cause significant UV noise in supercritical fluid chromatography (SFC). Deviations as little as 5 °C can increase noise as much as 5 times, making the detector unsuited for trace analysis. Two approaches were used to minimize this noise. When a flow cell was in direct thermal contact (metal on metal) with the detector optical bench, the mobile phase temperature was actively controlled to the measured flow cell temperature, by using one of the heat exchangers (HX) in the column compartment. However, with some older, but still widely used flow cell designs, this required repeated, hourly monitoring of the flow cell temperature and repeated manual adjustment of the heat exchanger temperature, due to thermal drift. Flow cell design had a strong influence on susceptibility to this thermally induced noise. Thermally insulating the flow cell from the optical bench made some cells much less susceptible to such thermally induced noise. Five different flow cells, some insulated, some un-insulated, were evaluated. Most had a truncated conical flow path, but one had a cylindrical flow path. Using either approach, the ASTM noise, with a 10mm, 13 μL conical flow cell, could be optimized to ≈0.007 mAU at 2.5 Hz, in SFC, which is very near the 0.006 mAU manufacturer's specification for HPLC. The insulated version of this flow cell required far less optimization, compared to the un-insulated version. At 150 bar, an experimental 3mm, 2 μL flow cell, with only one side insulated, yielded noise slightly too high (≈0.16-0.18 mAU) for trace analysis, at 80 Hz. However, at 200 bar, noise at 80 Hz was <0.06 mAU, which should allow quantification of a 1 mAU tall trace component with a signal to noise ratio (S/N) >10. Even partially un-insulated, this flow cell design was much less susceptible to thermally induced noise. Further insulating this flow cell design failed to improve performance.

Berger TA 《-》

Development of an orthogonal method for mometasone furoate impurity analysis using supercritical fluid chromatography.

While supercritical fluid chromatography (SFC) has received great popularity in chiral separation and purification, it has rarely been used for trace level pharmaceutical impurity analysis, partially due to the limitation of instrument sensitivity. In this study, a packed column SFC method has been developed for the quantitative analysis of mometasone furoate and its trace level impurities. The UV detection was optimized to improve the sensitivity by 2-4 fold. In combination with an increased sample concentration, this SFC method is capable of trace level (0.05% of the active) analysis of the impurities. The SFC method used a silica column and a mobile phase consisting of CO(2) and methanol. The new method provides an orthogonal selectivity complementary to the reversed phase HPLC (RP-HPLC) method. All of the impurities and the active were baseline separated within 12 min on SFC, which is less than one third of the RP-HPLC method run time. The method was also partially validated for linearity, accuracy, precision (repeatability), and limit of quantitation. This study demonstrated that the SFC method, with improved sensitivity, can be a valuable tool to provide orthogonal selectivity for trace level impurity separation. With further validation, the method may be suitable for release testing and stability testing for mometasone furoate drug substance.

Wang Z ，Zhang H ，Liu O ，Donovan B ... -  《-》

Simulated moving columns technique for enantioselective supercritical fluid chromatography.

Zhang Y ，Dai J ，Wang-Iverson DB ，Tymiak AA ... -  《CHIRALITY》

Direct coupling of packed column supercritical fluid chromatography to continuous corona discharge ion mobility spectrometry.

In this study, packed column supercritical fluid chromatography (SFC) was directly coupled to a continuous corona discharge (CD) ion mobility spectrometer (IMS) with several modifications. The main advantage of the developed detector is its capability to introduce full column effluent up to 2000 mL min(-1) CO(2) gas directly into the IMS cell relative to 40 mL min(-1) CO(2) gas as a maximum tolerance, reported for the previous IMS detectors. This achievement was made possible because of using corona discharge instead of (63)Ni as an ionization source and locating the inlet and outlet of the CO(2) gas in the counter electrode of the CD in opposite direction. In addition, a heated interface was placed between back pressure regulator (BPR) and the IMS cell to heat the output of the BPR for introducing sample as the gas phase into the IMS cell. Furthermore, a make-up methanol flow was introduced between the column outlet and BPR to provide a more uniform flow through the BPR and also to prevent freezing and deposition of the analytes in the BPR. The performance of the SFC-CD-IMS was evaluated by analysis of testosterone, medroxyprogesterone, caffeine, and theophylline as test compounds and figures of merit for these compounds have been calculated.

Rahmanian A ，Ghaziaskar HS ，Khayamian T 《-》