Polypyrrole as Adsorbent in Magnetic Solid Phase Extraction for Progesterone Determination from Human Plasma

Abstract

A straightforward and effective chromatographic method has been created for the analysis of progesterone from human plasma using a composite based on polypyrrole/magnetic nanoparticles in the sample preparation procedure. The quantification of progesterone is necessary due to its importance in human development and fertility. The employed conditions used acetonitrile:ultrapure water (70:30, v/v) as the mobile phase at 1.0 mL min^-1 and an octadecyl silane column (Phenomenex Gemini, 250 mm × 4.6 mm, 5 μm) at a wavelength of 235 nm. The composite and its precursors were synthesized and properly characterized by X-ray diffraction, Fourier transform infrared spectroscopy, scanning electron microscopy/energy dispersive spectroscopy, thermogravimetric analysis, and point of zero charge. The main factors affecting the extraction recovery of progesterone from pool human plasma samples employing magnetic solid phase extraction have been studied, such as sample pH (without adjustment), sample volume (1000 μL), washing solvent (ultrapure water), eluent (acetonitrile), eluent volume (1000 μL), and amount of adsorbent (10 mg). The extraction recoveries ranged from 98% to 102%, and linearity ranged from 5 to 3000 ng mL^-1. The correlation coefficient (r) was ≥0.99, and acceptable relative standard deviation (precision), relative error (accuracy), and p-values (robustness) were observed. Lastly, the plasma samples from pregnant women were successfully analyzed by the validated method.

Figure 1

(A) TGA, (B) FTIR, and (C) XRD data of Fe₃O_4, Fe₃O₄@SiO₂, and MMPPy. (D) BET nitrogen adsorption–desorption isotherm plot of MMPPy.

Figure 2

SEM images at magnifications of 500× and 2000× for (A, B) Fe₃O₄, (C, D) Fe₃O₄@SiO₂, and (E, F) MMPPy.

Figure 3

Sample preparation optimization using MMPPy in MSPE: (A) washing solvent, (B) washing solvent volume, (C) eluent type, (D) eluent volume, (E) amount of adsorbent, (F) sample volume, and (G) sample pH. (H) Graphical results from the AGREEprep analysis calculated by emulator version online.

Figure 4

(A) Reuse (1–4×) and (B) comparison of different materials: MMPPy, Fe₃O₄@SiO₂PAni, and Fe₃O₄@PAni mesoporous.

Figure 5

Chromatograms referring to the determination of (A) progesterone (P4) standard at 10 μg mL^–1, (B) spiked human plasma sample at 10 μg mL^–1 with P4 (green line) and blank pool human plasma sample (blue line), (C) 35 weeks pregnant woman plasma, and (D) 8 weeks pregnant woman plasma. Chromatographic conditions: Phenomenex Gemini C18 analytical column (250 mm × 4.6 mm, 5 μm) without temperature control, mobile phase composed of acetonitrile:ultrapure water (70:30, v/v), flow rate at 1.0 mL min^–1, 20 μL injection volume, and detection at λ = 235 nm.