UCT has produced an application note outlining a suggested procedure that reduces column backpressure while also reducing analyte suppression, when compared to a standard dilute and shoot protocol. Using UCT’s Refine™ Ultra-Filtration Technology allows for sample precipitation and filtration to occur simultaneously.
Introduction
Various biological sample types commonly contain proteins that complicate downstream analysis if not effectively removed. While sample preparation techniques, such as solid-phase extraction (SPE), can successfully remove these unwanted matrix components, complex and lengthy extraction procedures are not ideal for high-throughput laboratories. With the advancement of LC-MS/MS technology, many laboratories have implemented dilute and shoot procedures in order to reduce sample preparation time, reduce consumable spending and reduce overall turnaround time. The caveat to this quick protocol is the toll that it ultimately takes on the HPLC column and instrument source.
Using UCT’s Refine™ Ultra-Filtration Technology a novel, hydrophobically-treated, submicron frit combination facilitates the removal of sample proteins without a complicated extraction. Column backpressure is reduced in addition to analyte suppression, when compared to a standard dilute and shoot protocol.
Results - Analyte Suppression
Upon passing urine samples containing hydrolysis enzyme through the Refine™ Ultra-Filtration plate, analyte suppression was significantly reduced across the board for the monitored benzodiazepine and opiates panel ranging from 7-71% improvement. Samples were done in triplicate.
Results – Backpressure Stabilization
Backpressure per sample injection was also assessed when comparing the utilization of Refine™ Ultra-Filtration technology versus a Dilute & Shoot (D&S) preparation for hydrolyzed urine samples. After 50+ injections, the backpressure remained stable when utilizing the Refine™ plate as opposed to D&S, where a drastic increase was seen after as little as 10 injections.
Conclusion
Using the above procedure, the Refine Ultra-Filtration technology allowed for a reduction in instrumental backpressure and removal of unwanted matrix components in both urine and serum samples. Overall, this allows for an end user to have enhanced analyte selectivity and an increase in HPLC/UHPLC column life. The extraction procedure can be completed in approximately 20 minutes and can easily replace a dilute and shoot protocol with minimal additional investment in laboratory resources. The hydrophobically-treated submicron frit combination allows for efficient sample protein precipitation and filtration and is amenable to both forensic and clinical laboratory settings.
