Automated Solutions using Small Scale SPE to Solve Challenging Applications

Importance of the Topic

The automation of small-scale solid phase extraction (SPE) addresses critical challenges in high-throughput laboratories. By integrating Instrument Top Sample Preparation (ITSP) on autosamplers, method reproducibility and sample throughput increase significantly. This approach is vital for environmental monitoring of trace contaminants, clinical assays requiring consistent bioanalysis, and food and flavor quality control where low detection limits and robust sample cleanup are mandatory.

Objectives and Study Overview

This technical note summarizes multiple automated ITSP workflows developed by Anatune Ltd. over eight years. The primary goals were to demonstrate the versatility of ITSP for a range of analytes—vitamin D metabolites in human serum, N-Nitrosodimethylamine (NDMA) and metaldehyde in water, taste and odour compounds, and filtration strategies for UHPLC—and to showcase system robustness and throughput across environmental, food/flavor, and clinical applications.

Methodology and Sample Preparation

The core ITSP workflow employs a Gerstel MPS 2 XL autosampler equipped with dual heads: a 2.5 mL headspace syringe for SPE operations and a 10 µL syringe for injection. Sample preparation steps include:

• Protein precipitation and internal standard addition for serum samples targeting 25-OH D2 and D3, followed by magnetic transport to a CF100 centrifuge for vortexing and centrifugation.
• Conditioning of C18 ITSP cartridges, loading of supernatant, washing, drying with air, and elution with methanol for direct LC–MS/MS injection.
• Extraction of NDMA and metaldehyde from water using coconut charcoal or ENV cartridges, with critical nitrogen drying steps, followed by elution into GC vials and GC–MS/MS analysis.
• SPE enrichment of taste and odour compounds from 35 mL water samples spiked with internal standards, salt addition, equilibration, cartridge drying, elution, and GC–MS/MS measurement.
• Filtration of samples through ITSP cartridges with acetone elution to protect UHPLC systems from particulates.

Instrumentation

• Gerstel MultiPurpose Sampler (MPS) 2 XL with ITSP tray
• Anatune CF100 centrifuge and agitator
• Agilent 1200 LC coupled to 6410 Triple Quadrupole MS/MS using APCI
• Agilent 6460 MS/MS at Guy’s and St Thomas’ Hospital for vitamin D analysis
• Agilent 7000 GC Triple Quadrupole with EI source and CIS 4 PTV cooled injection system

Main Results and Discussion

Vitamin D assays achieved linear calibrations (R2 ≥ 0.998) over 3.8–37.5 µg/L for 25-OH D2 and 25-OH D3, with weekly throughput exceeding 200 serum samples. NDMA extraction delivered linearity (R2 = 0.9996) from 0.25 to 15.25 ng/L. Metaldehyde calibration from 0.04 to 0.8 µg/L showed excellent linearity and reproducibility. Taste and odour compounds exhibited low detection limits (0.6–1.1 ng/L) and high coefficients of determination (R2 > 0.99). System robustness was demonstrated by routine analysis (up to 100 water samples weekly) with minimal manual intervention.

Benefits and Practical Applications

Automated ITSP offers:

• Enhanced reproducibility through standardized conditioning, loading, and elution steps.
• Higher sample throughput and reduced hands-on time.
• Improved laboratory safety by limiting solvent handling.
• Scalable workflows adaptable to clinical diagnostics, environmental compliance, and food safety testing.

Future Trends and Applications

Advancements in miniaturized SPE cartridges and integration with ultrahigh-resolution mass spectrometry are expected to lower detection limits further. Coupling ITSP with online sample derivatization and real-time data processing will expand its use in metabolomics and rapid screening for emerging contaminants. Automation platforms may evolve to support multi-dimensional separations and direct coupling with ambient ionization sources.

Conclusion

The ITSP-based workflows reviewed here demonstrate a robust, versatile platform for automated SPE in diverse analytical contexts. Their proven linearity, sensitivity, and throughput underscore the value of integrating sample preparation with autosampler systems to meet the demands of modern analytical laboratories.