Rp 18 60 f254

The following Standards were used in the study: sildenafil citrate (SLD) (Purity > 99%) (Fig. 1) (Sigma-Aldrich, St. Louis, MO, USA), SLD (Viagra) (Pfizer Inc, USA), hydroxyprogesterone (Purity ≥ 99%) (Bayer Pharma AG, Germany) and oestradiol benzoate (Purity ≥ 98%) (Misr Co. for Pharm. Ind., Egypt). Four marketed products (Film coated Tablet, 50 mg) approved in Saudi Arabia were obtained from Drug stores in Riyadh. Chromatographic and analytical grade reagent (AR) were used for extraction and method development. Pre-coated, glass-baked TLC plates obtained from E. Merck (RP-18-60F254; thickness: 0.2 mm; area: 20 × 10 cm) were used for quantification.Figure 1

Structures of Sildenafil (SLD) and Sildenafil N-oxide (SLD N-oxide).

HR-ESI-MS and ESI-MS were obtained with a Bruker microTOFQ mass spectrometer (Bruker Daltonics, Bremen, Germany). The NMR spectral data were recorded on a Bruker AV-500 FT-NMR (500 MHz for ¹H and 125 MHz for ¹³C) in C₅D₅N, using visual C₅D₅N resonances (δ_H 7.21, 7.58, and 8.73, δ_C 123.5, 135.5, and 149.0) for internal reference. All chemical shifts (δ) are given in ppm. Optical rotations were measured by using a JASCO P-1020 automatic digital polarimeter (JASCO Corporation, Tokyo, Japan). Preparative HPLC was performed on a NP7005C pump connected with a SHODEX RI-102 detector (Shoko Scientific, Tokohama, Japan), using Megres ODS column (250 mm × 20 mm, i.d., 5 μm, Hanbang Sci. and Tech., Haian, China). Column chromatography was performed with macroporous resin HPD100 (Cangzhou Bon Adsorber Technology, Cangzhou, China) and RP-18 reversed-phase silica gel (S-50 mm, YMC, Kyoto, Japan). TLC analysis was carried out on pre-coated TLC plates with silica gel RP-18 60 F₂₅₄ (Merck, Darmstadt, Germany, 0.25 mm). Detection was achieved by spraying with 10% H₂SO₄ in MeOH followed by heating. HPLC-grade MeOH was purchased from Merck. HPLC-grade water was purified using a Milli-Q system (millipore, Boston, MA, USA). All solvents used for the chromatographic separations were distilled before use.

A Bruker AV-500 FT-NMR spectrometer was used, operating at 500.1 MHz for ¹H and at 125.8 MHz for ¹³C, chemical shifts are expressed in δ referring to the residual solvent signals δ_H 2.50 and δ_C 39.5 for DMSO-d₆, coupling constants, J, are in hertz. ESI-MS was acquired with a Bruck micro-TOFQ mass spectrometer (Bruck, Bremen, Germany). Column chromatography was performed over a RP-18 reversed-phase silica gel (S-50 μm; YMC, Kyoto, Japan). Thin-layer chromatography (TLC) analysis was carried out on a pre-coated TLC plate with silica gel RP-18 60 F254 (0.25 mm, Merck, Darmstadt, Germany). Detection was achieved by spraying the sample with 10% H₂SO₄ in MeOH followed by heating. Preparative HPLC was performed using a Shimadzu LC-6AD pump connected to a Shimadzu SPD-20A UV-VIS detector (at 254 nm) with a Shim Pak ODS column (250 mm × 21.2 mm, i.d., 10 μm, Shimadzu, Kyoto, Japan) and a Megress ODS column (250 mm × 20 mm, i.d., 10 μm, Jiangsu hanbon Science and Technology Co., Ltd., Huai′an, China). Analytical HPLC was performed with two LC-20AT solvent delivery pumps, an online mixer, a SIL-20A autosampler, a CTO-20A column temperature controller, and a SPD-M20A photodiode-array detector (Shimadzu). HPLC-grade water was purified using a Milli-Q system (Millipore, Boston, MA, USA). All solvents used for the chromatographic separations were distilled before use.