Sample Preparation, HPLC
IndustriesManufacturerKNAUER
Importance of the Topic
Polycyclic aromatic hydrocarbons are ubiquitous environmental contaminants with significant mutagenic and carcinogenic properties. Sensitive and reliable analytical methods are essential to detect trace levels in complex matrices. Online coupling of solid phase extraction with high performance liquid chromatography offers automated sample processing, matrix reduction and enhanced sensitivity, making it highly valuable in environmental monitoring and quality control.
Objectives and Study Overview
This study illustrates a versatile online SPE–HPLC setup employing an eight-port two-position valve and a multi-injection valve for the analysis of 16 PAHs according to EPA method 610. The configuration enables switching between manual loading, autosampler injection and use of an auxiliary feed pump without reconfiguring the plumbing.
Methodology
The EPA 610 PAH standard was diluted in acetonitrile/water (60:40 v/v) at various concentrations. Three valve modules provided multiple flow paths: manual syringe loading via the multi-injection valve, automated loading via autosampler, and pump-driven loading through an auxiliary pump. Valve switching sequences governed cartridge conditioning, sample loading, elution and cleaning. Elution was performed with a binary gradient pump, directing analytes onto a C18 main column for separation and detection by UV and fluorescence.
Used Instrumentation
- Binary HPLC Pump AZURA P 6.1L HPG, 10 ml, 862 bar
- Autosampler AZURA AS 6.1L
- DAD Detector AZURA DAD 2.1L, UV at 230 nm
- Fluorescence Detector RF-20A
- Temperature Controller AZURA CT 2.1
- Valve Unifier VU 4.1, multi-position valve 8 port 1/8'' and high-pressure injection valve 8 port 2-position
- Multi-injection Valve DLC stainless steel
- SPE Cartridge Bioselect 300-5 C8, 30×4 mm
- Analytical Column NUCLEOSIL 100-5 C18 PAH, 150×4 mm, 5 µm
- ClarityChrom 8.5 Software with autosampler and PDA control
Main Results and Discussion
Chromatographic separation of all 16 target PAHs was achieved with distinct peaks in UV and fluorescence detection modes. Automated SPE loading via the auxiliary pump provided the highest sample throughput and sensitivity, while autosampler loading offered flexibility for small volumes. Manual loading demonstrated feasibility but was limited by cartridge backpressure. Optimized valve sequences ensured reproducible conditioning, loading and elution cycles.
Benefits and Practical Applications
- Automation reduces hands-on time and risk of error
- Matrix cleanup improves detection limits
- Flexible loading options accommodate sample volumes from microliters to milliliters
- Modular valve setup allows rapid method adaptation
Future Trends and Potential Applications
Integration of online SPE with advanced detectors and mass spectrometry will further enhance selectivity and sensitivity. Miniaturized cartridge formats and high-throughput valve arrays may allow parallel sample processing. Applications could extend to pharmaceuticals, food safety and complex biological matrices.
Conclusion
The presented online SPE–HPLC method with multi-position and multi-injection valves offers a robust and adaptable platform for trace analysis of PAHs. It combines automation, sensitivity and methodological flexibility, making it suitable for routine environmental and industrial monitoring.
References
- NCBI PMC7674206 Article on global occurrence of PAHs, accessed 2022-03-21
- EPA Method 610 for PAH analysis, published 1984, accessed 2022-03-21
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