A Forensic Investigation into the Presence of Phosphodiesterase 5 Inhibitors as Adulterants in Herbal Remedies
- Publication Type:
- Thesis
- Issue Date:
- 2020
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The proliferation of herbal-based male sexual performance products, particularly those adulterated with phosphodiesterase 5 (PDE5) inhibitors, has sparked grave public health and food safety concerns. The advent of their unapproved analogues presents an additional challenge to forensic drug testing laboratories, as these adulterants may evade detection during routine screening. A comprehensive strategy is warranted to address these problems and protect consumers’ health and well-being. This study investigated the presence of PDE5 inhibitors as adulterants in herbal remedies, using a two-tier screening strategy of rapid qualitative assay and confirmatory analytical analysis.
A bioactivity-based PDE5 inhibition assay was established using fluorescein-labelled cyclic-3’,5’-guanosine monophosphate as substrates to PDE5 enzyme. The PDE5 inhibitions, measured using a fluorescence polarisation technique, was applied to 50 herbal-based food samples. The results were in agreement with the confirmatory analytical analysis for all food products, except for the instant coffee premix samples, postulated due to the presence of caffeine. The assay, nevertheless, exhibited a promising potential to rapidly screen PDE5 inhibitors in various types of food products, except those containing naturally-occurring phosphodiesterase inhibitors.
A confirmatory liquid chromatography-high-resolution mass spectrometry (LC‑HRMS) analysis was developed using 23 target analytes; selected to represent different groups of PDE5 inhibitors, based on their structural similarities. The targeted analysis was primarily optimised to mitigate the matrix effect (ME), via chromatographic separation, sample extraction, and sample dilution. The insignificant ME percentages, within ‑9.2%–8.8% for all target analytes in food and pharmaceutical matrices, were evidenced with satisfactory validation results; notably, the accuracy was within 77.4%–124.7%. The development, optimisation, and validation of the targeted analysis provided a solid foundation for suspected-target and non-targeted screenings. The suspected-target screening employed a library comprising 95 PDE5 inhibitors, providing extended coverage of known analytes. Contrarily, the non-targeted screening adopted top-down and bottom-up approaches to flag novel PDE5 inhibitors analogues based on common fragmentation patterns of target analytes.
The confirmatory LC-HRMS analysis was applied to 50 herbal-based food samples and 52 herbal-based pharmaceutical samples. The targeted analysis and the suspected-target screening identified 11 target analytes and detected five suspected analytes, respectively, from 74 adulterated samples. The non-targeted screening returned insignificant signals, indicating the absence of potentially novel analogues. Some of these samples contained up to five different PDE5 inhibitors and quantified at supratherapeutic level, making them unsafe for consumption. The comprehensive strategies provide a superior approach to curb the widespread adulteration of herbal remedies, thus, safeguarding the public’s health.
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