title: Validated UPLC-MS/MS assay for the determination of synthetic phosphodiesterase type-5 inhibitors in postmortem blood samples.
abstract: The use of synthetic phosphodiesterase type 5 (PDE-5) inhibitors for the treatment of erectile dysfunction: sildenafil citrate (Viagra(®)), tadalafil (Cialis(®)) and vardenafil hydrochloride (Levitra(®)) has increased dramatically over the past 2 years. These substances are prescription drugs and must be used under medical supervision. However, they can easily be obtained over the internet from illegal sites, being a potential for a threat to public health. The development of an electrospray ionisation (ESI) ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) procedure for the simultaneous identification and quantification of three PDE5 inhibitors in blood samples was desired. Samples were prepared using Oasis(®) HLB solid-phase cartridges (3 cc, 60 mg) and chromatographic separation was achieved on an Acquity UPLC(®) HSS T3 (100 × 2.1 mm i.d., 1.8 μm particles) column with a gradient mobile phase of 0.1% formic acid and acetonitrile at a 0.5 mL/min flow rate. Quantification was achieved by multiple reaction monitoring (MRM) of two transitions per compound: m/z 475.1 > 58 e m/z 475.1 > 311.1 for sildenafil; m/z 389.9 > 267.9 e m/z 389.9 > 134.8 for tadalafil and m/z 489 > 71.9 e m/z 489 > 150.9 for vardenafil. Zolpidem-d6 (m/z 314.5 > 235.3) was used as the internal standard. Calibration curves were linear over the concentration range of 5-1000 ng/mL, with a coefficient of determination better than 0.997. The lower limits of detection and quantification for these substances were ≤ 3 ng/mL and ≤ 8 ng/mL, respectively. The method showed a satisfactory sensitivity, precision, accuracy, recovery and selectivity. A rapid, selective and sensitive UPLC-MS/MS method using solid-phase extraction was developed for the simultaneous determination and quantification of sildenafil, vardenafil and tadalafil in blood samples.
authors.affiliation: National Institute of Legal Medicine, Largo da Sé Nova, 3000-213 Coimbra, Portugal. paulaproenca@dcinml.mj.pt
authors.forename: Paula
authors.initials: P
authors.lastname: Proença
authors.forename: Carla
authors.initials: C
authors.lastname: Mustra
authors.forename: Mariana
authors.initials: M
authors.lastname: Marcos
authors.forename: João Miguel
authors.initials: JM
authors.lastname: Franco
authors.forename: Francisco
authors.initials: F
authors.lastname: Corte-Real
authors.forename: Duarte Nuno
authors.initials: DN
authors.lastname: Vieira
mesh_terms: D002243:Carbolines / Q000097:blood; D002853:Chromatography, Liquid; D053593:Forensic Toxicology / Q000379:methods; D006801:Humans; D007093:Imidazoles / Q000097:blood; D057230:Limit of Detection; D008297:Male; D058986:Phosphodiesterase 5 Inhibitors / Q000097:blood*; D010879:Piperazines / Q000097:blood; D011687:Purines / Q000097:blood; D000068677:Sildenafil Citrate; D021241:Spectrometry, Mass, Electrospray Ionization; D013450:Sulfones / Q000097:blood; D000068581:Tadalafil; D053719:Tandem Mass Spectrometry; D014227:Triazines / Q000097:blood; D000069058:Vardenafil Dihydrochloride
chemical_list: D002243:Carbolines; D007093:Imidazoles; D058986:Phosphodiesterase 5 Inhibitors; D010879:Piperazines; D011687:Purines; D013450:Sulfones; D014227:Triazines; D000069058:Vardenafil Dihydrochloride; D000068581:Tadalafil; D000068677:Sildenafil Citrate
keywords: Sildenafil; Tadalafil; UPLC-MS/MS; Vardenafil