Portable liquid chromatography for on-site analysis
Creators: name: Hemida, MHS
Publisher: name: University Of Tasmania
Year: 2024

description: High performance liquid chromatography (HPLC) is a dominant laboratory-based analytical technique, but despite its dominant position within analytical laboratories, HPLC has not seen significant application to-date in many important field-based applications. For example, a portable and field-deployable liquid chromatographyultraviolet/ mass spectrometric system (LC-UV/MS) would be of considerable value to facilitate on-site analysis for the pharmaceutical manufacturing sector, areas of industrial process monitoring, threat analysis, and environmental monitoring. However, there are major challenges associated with development of fully functional and field-deployable portable LC-UV/MS systems, particularly for the pharmaceutical manufacturing sector and related on-site industrial applications. Therefore the main aims of this research have been to; (1) Develop and demonstrate the analytical performance of miniaturized UV absorbance detectors with suitability for application within portable capillary LC systems; (2) Undertake a configuration and compatibility investigation into the coupling of a portable briefcase-sized capillary LC-UV platform with new portable mass spectrometric (MS) detector for point-of-need LC-UV-MS analysis; (3) Integrate and demonstrate the performance of the portable LC with industry-standard auto-sampling devices for automated process analysis within pharmaceutical laboratories; and (4) Undertake an in-field demonstration of the coupled portable LC/MS system for environmental analysis. Light-emitting diodes (LEDs) are small in size, have low power consumption requirements, short warm-up times, and deliver long lifetimes. These make LEDs attractive light sources for the miniaturization of absorbance-based detectors. However, LEDs are quasi-monochromatic with narrow emission spectra. This significantly limits the potential of LEDs for applications that require multi-wavelength detection capability. A number of limitations and improvements in low-UV LED-based absorbance detectors were investigated within the first phase of this research. A new miniature deep multi- LED UV absorbance detector was developed using low-cost LEDs, which could be operated in both individual wavelength (240, 255, and 275 nm) and scanning (230 - 300 nm) detection modes. The detector was mostly composed of off-the-shelf components, such as the LEDs, a trifurcated fiber assembly (TFFA), a commercial capillary Z-type flow cell, and the detector photodiodes. The detector was subsequently incorporated within a portable 'briefcase-sized' capillary LC system. The detector was characterized in terms of performance and was further benchmarked against a commercial variable wavelength capillary LC detector (Dionex UltiMate 3000RS). The developed detector showed low levels of stray light (&lt;0.4%), delivered an effective path length of up to 99.0% of the detection flow cell, gave a wide dynamic range (0.5 to 200 μg/mLfor sulfamethazine, carbamazepine, and flavone), and exhibited low noise levels (at 300 μAU level). Further to the above, in the second phase of this research, the design and performance demonstration of a compact, modular and broadband (D<sub>2</sub> lamp-based) UV-Vis absorbance detector was investigated, as an alternative to the LED-based detectors. The detector was composed of a miniature D<sub>2</sub> light source, an end-capillary extended optical pathlength (10 mm) flow-cell, and a miniature spectrometer. The assembled detector showed low levels of stray light (&lt;0.2%), utilization of up to 91.0% of the effective path length of the Z-type flow-cell, a linearity limit of ~ 3.0 AU, and low noise levels (58.0×10<sup>-5</sup> AU). Further, the detector was again integrated and demonstrated with the above portable capillary LC and was then also benchmarked against a commercial capillary LC detector. The detector provided full UV-Vis scanning spectral capability for analytes when used with the portable capillary HPLC. Limits of detection (LODs) for five test pharmaceuticals ranged from 190 to 602 μg L<sup>−1</sup>. The system was then demonstrated for applications within the pharmaceutical manufacturing sector. Integrating a portable LC system as a process analytical technology (PAT) within industrial settings is a significant technical challenge. This typically includes several steps, including automated sampling, automated sample preparation, the actual chromatographic analysis, and subsequent data processing. In the third phase of this research, the portable capillary LC with the above miniaturized detection options was deployed as a PAT system within a commercial pharmaceutical laboratory. The system was integrated with an industry standard automated sampling device and used for studying the kinetics of a synthetic reaction and benchmarked against a commercial process laboratory full ultra-high performance LC system. The deployed system gave low detection limits (3 μg L<sup>−1</sup>) and delivered a wide dynamic range ((up to 200 μg mL<sup> −1</sup>). The on-site demonstration confirmed robust performance in automated process analysis, with less than 5.0% relative standard deviation (RSD) on sample-to-sample reproducibility, and less than 2% carry-over between samples. The system has shown potential to significantly increase sample throughput by providing near real-time analysis and to improve understanding of synthetic processes. Further to the above industrial application, it should be noted that there are very few reports to-date of field-deployable LC-based systems which can be successfully employed for on-site trace analysis. However, portable and field-deployable LC-MS systems for in-field analysis of complex mixtures and highly challenging samples is a definite future need. In the final phase of this research work, the portable and field-deployable capillary LC-MS system was configured for in-field analysis of per- and polyfluoroalkyl substances (PFAS) in soils. Thus, a portable ultrasound-assisted extraction (UAE) methodology was developed, optimized, and applied with the portable capillary LC/MS system for on-site analysis of PFAS in real soil samples. The critical parameters for the integration of the capillary LC with MS detection, and on the separation and performance of the coupled capillary LC/MS were studied and optimized. The UAE method were also studied and optimized using response surface methodology (RSM) based on a central composite design (CCD). Spiked soil samples with known concentrations of 50 µg/L or 100 µg/L were used for method validation. The mean recovery for eleven PFAS ranged between 70 - 110%, with relative standard deviations (RSD) ranging from 3 to 12%. When deployed to a field site, method reporting limit for 12 PFAS ranged from0.6 to 0.1 ng g <sup>-1</sup>, with wide dynamic ranges (1 - 600 ng g<sup> -1</sup>) and excellent linearities (R<sup>2</sup> ˃ 0.991). The in-field portable system was again benchmarked against a commercial lab-based LC/MS for the analysis of PFAS in real soil samples, with comparison results showing good agreement. Twelve PFAS were detected and identified in real soil samples at concentrations ranging from 8.1 ng g <sup>-1</sup>) (for perfluorooctanesulfonic acid, PFOS) to 2.9 µg g <sup>-1</sup> (perfluorohexanesulphonic acid, PFHxS). The system demonstrated significant potential and robustness, and it is anticipated the system could help to deliver sensitive and fast analysis for rapid on-site environmental monitoring in the near future.