Extraction of n-Hexane-Extractable Material from Wastewater for U.S. EPA Method 1664B with the Biotage VacMaster Disk, Using 90 mm SPE Disks

Significance of the topic

Accurate determination of n-hexane-extractable material (HEM) in wastewater is essential for environmental monitoring and regulatory compliance. EPA Method 1664B defines standardized procedures for quantifying oils and greases; implementing a robust extraction system improves data quality, throughput and laboratory efficiency.

Study objectives and overview

This application note evaluates the performance of the Biotage VacMaster Disk system for HEM extraction from 1 L wastewater samples according to EPA Method 1664B. The study established the minimum level of quantitation (ML), method detection limit (MDL) and initial precision and recovery (IPR) to demonstrate system capability.

Methodology and instrumentation

Sample pre-treatment and extraction steps were conducted on eight VacMaster Disk stations daisy-chained to a single vacuum pump. Key steps:

• Acidify samples to pH < 2 using HCl.
• Fortify blanks and reagent water with hexadecane and stearic acid spikes at defined concentrations.
• Condition 90 mm O&G SPE disks with n-hexane and methanol before loading.
• Load 1 L sample under vacuum, dry disk, then elute analytes with three sequential n-hexane rinses (total 70 mL).
• Dry extracts over anhydrous sodium sulfate, evaporate solvent by SpeedVap IV at 40 °C, weigh residue to report mg/L HEM.

Instrumentation used:

• Biotage VacMaster Disk system with 90 mm reusable disk holders.
• Pacific Premium O&G SPE disks (O.D. 90 mm).
• Vacuum pump set to –24 in Hg.
• SpeedVap IV solvent evaporation system.

Main results and discussion

ML study (n = 4 blanks) yielded an average background HEM of 0.3 mg/L, well below the 5.0 mg/L acceptance criterion. The MDL study (n = 7 spikes at 4 mg/L) produced a calculated MDL of 0.47 mg/L versus a regulatory limit of 1.4 mg/L. IPR (n = 4 reagent water replicates spiked at 20 mg/L) delivered average recoveries of 89.6% (range 87–93%) and a standard deviation of 2.6%, meeting the EPA criteria (83–101% recovery and ≤ 11% RSD).

Benefits and practical applications

The VacMaster Disk approach provides:

• High throughput: simultaneous processing of up to eight samples.
• Reproducible low-level quantitation suitable for regulatory compliance.
• Compact footprint and simplified workflow compared to conventional liquid–liquid extraction.
• Reduced solvent consumption and waste handling.

Future trends and opportunities

Automation and integration with in-line detection could further streamline HEM analysis. Adaptation of disk-based cleanup for broader analyte classes, use of greener solvents, and digital tracking of sample workflow represent promising developments. Expanded data connectivity will enable real-time QA/QC monitoring and method validation across multiple laboratories.

Conclusion

The Biotage VacMaster Disk system delivers compliant, precise and efficient extraction of n-hexane-extractable material in wastewater per EPA Method 1664B. Its scalability, reproducibility and low detection limits support routine environmental monitoring and quality assurance programs.

Reference

• EPA Method 1664, Revision B, February 2010.
• 40 CFR Part 136, Appendix B – Definition and Procedure for the Method Detection Limit, Revision 1.11.