Detection and Identification of Synthetic Phosphodiesterase Type-5 Inhibitors in Adulterated Herbal Supplements Using UPLC and Data-Directed Analysis by Mass Spectrometry

Importance of the topic

Herbal dietary supplements promoted as natural remedies for erectile dysfunction have been frequently discovered to contain undeclared synthetic PDE5 inhibitors. These adulterants pose significant health risks because consumers may unknowingly ingest potent pharmaceuticals without medical supervision. Rapid and reliable analytical methods are essential to protect public health by detecting both known compounds and emerging analogues in complex supplement matrices.

Objectives and overview of the study

This work evaluates an integrated UPLC–tandem quadrupole mass spectrometry approach using the Xevo TQ MS with data-directed acquisition for the detection, identification, and quantitation of synthetic PDE5 inhibitors in herbal supplement samples purchased over the Internet. The aim is to demonstrate a single-run screening strategy that acquires both MS and MS/MS data in real time, improving throughput and structural elucidation capability.

Used methodology and instrumentation

Samples of tablets and capsules were finely ground and extracted in 50:50 methanol/water by sonication. After centrifugation, extracts were injected (5 μL) onto an ACQUITY UPLC BEH C18 column (2.1 × 100 mm, 1.7 μm) maintained at 60 °C, using a gradient of ammonium acetate buffer (pH 6.7) and methanol/acetonitrile. Flow rate was 550 μL/min.
Chromatography was coupled to a Xevo TQ MS operated in positive ESI. The instrument performed high-speed survey scans (up to 5000 amu/sec) and automatically switched to MS/MS (ScanWave DS) upon peak detection. Collision energies of 20–40 eV were used to generate diagnostic fragment ions for analyte confirmation.

Used instrumentation

• ACQUITY UPLC system with Binary Solvent Manager and Sample Manager
• ACQUITY UPLC BEH C18 column (2.1 × 100 mm, 1.7 μm)
• Xevo TQ MS tandem quadrupole mass spectrometer with ScanWave collision cell
• Electrospray ionization source operated in positive mode

Main results and discussion

Five herbal products were analyzed and all were found to be adulterated:

• Sample 1: contained only tadalafil
• Sample 2: contained sildenafil
• Sample 3: contained both sildenafil and tadalafil plus additional unknown analogues
• Sample 4: co-adulterated with sildenafil and tadalafil
• Sample 5: lacked reference standards but yielded MS/MS fragments indicative of sildenafil analogues

Data-directed acquisition allowed simultaneous MS and MS/MS data collection, enabling rapid confirmation of active ingredients by their characteristic fragment ions. Quantitation (external calibration 0.1–500 ng/mL) revealed sildenafil levels ranging from 0.078 to 116.7 mg per dose and tadalafil levels of 13.5 to 52.1 mg per dose, comparable to prescription doses and thus presenting potential overdose risks.

Benefits and practical applications of the method

• Single-run screening eliminates separate full-scan and targeted MS/MS injections
• Real-time switching improves productivity and reduces sample consumption
• High scan speed ensures adequate peak definition for UPLC narrow peaks
• Capability to detect unknown analogues through common fragment or neutral-loss scanning
• Quantitative performance suitable for regulatory and quality-control laboratories

Future trends and potential applications

Further developments may include expansion of data-directed libraries for rapid analogue identification, incorporation of high-resolution mass spectrometry for exact mass determination, and automated data processing workflows to enable real-time surveillance of online supplement markets. Portable or field-deployable versions could support on-site screening by regulatory agencies.

Conclusion

The combination of UPLC with the Xevo TQ MS and data-directed acquisition provides a robust, high-throughput platform for the detection, identification, and quantitation of synthetic PDE5 inhibitors in adulterated herbal supplements. This approach enhances public health protection by enabling rapid screening of both known drugs and emerging analogues in a single analytical run.

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