Mu Yuan

Pharmacokinetics, tissue distribution and relative bioavailability of puerarin solid lipid nanoparticles following oral administration

Puerarin has various pharmacological effects; however, poor water-solubility and low oral bioavailability limit its clinical utility. A delivery system of solid lipid nanoparticles could enhance its oral absorption. The objective of this study was to investigate the pharmacokinetics, tissue distribution and relative bioavailability of puerarin in rats after a single dose intragastric administration of puerarin solid lipid nanoparticles (Pue-SLNs). The puerarin concentrations in plasma and tissues were determined by rapid resolution liquid chromatography electrospray ionization-tandem mass spectrometry. The C(max) value of puerarin after the administration of Pue-SLNs was significantly higher than that obtained with puerarin suspension (0.33±0.05 μg/mL vs. 0.16±0.06 μg/mL, P<0.01). The T(max) value after the administration of the Pue-SLNs was significantly shorter than that after puerarin suspension administration (40±0 min vs. 110±15.49 min, P<0.01). The AUC(0→t) values of puerarin were 0.80±0.23 mg h/L, and 2.48±0.30 mg h/L after administration of the puerarin suspension and Pue-SLNs, respectively. Following administration of the Pue-SLNs, tissue concentrations of puerarin also increased, especially in the target organs such as the heart and brain. These data suggest that SLNs are a promising delivery system to enhance the oral bioavailability of puerarin.

Determination of puerarin in rat plasma by rapid resolution liquid chromatography tandem mass spectrometry in positive ionization mode

A highly sensitive and specific method of rapid resolution liquid chromatography tandem mass spectrometry (RRLC-MS/MS) in positive ionization mode has been developed and validated for pharmacokinetic study of puerarin in rat plasma. Chromatography was carried out on a Zorbax XDB C18 reversed-phase column using a mobile phase comprising a mixture of methanol and 0.05% acetic acid in water (35:65, v/v) with a flow rate of 0.3 mL/min from 0 min to 5.4 min and then 0.6 mL/min from 5.41 min to 12 min. The mass spectrometer operated in ESI positive ionization mode. Multiple reaction monitoring (MRM) was used to measure puerarin and tectoridin (internal standard). The method was sensitive with a detection limit of 0.33 ng/mL. A good linear response was observed over a range of 10-2000 ng/mL in rat plasma. The inter- and intra-day precision ranged from 2.97% to 7.52% and accuracy from 93.70% to 101.60%. This validated method was applied successfully to a pharmacokinetic study in rat plasma after intravenous administration of puerarin. The main pharmacokinetic parameters were as follows: AUC(0→t) 45.37±13.19 (mgh/L), AUC(0→∞) 47.03±14.78 (mgh/L), MRT 1.03±0.46 (h), T(1/2) 1.31±0.31 (h), V(ss) 0.09±0.02 (L), V(z) 0.17±0.04 (L), Cl 0.10±0.04 (L/h).

Ligand-based pharmacophore model of N-Aryl and N-Heteroaryl piperazine α1A-adrenoceptors antagonists using GALAHAD

Computer aided drug discovery for selective antagonism effects on alpha(1A) subtypes of G-protein coupled receptors are important in the treatment of benign prostatic hyperplasia (BPH). Ligand-based pharmacophore models of N-Aryl and N-Heteroaryl piperazine alpha(1A)-antagonists were developed using two separate training sets. Pharmacophore models were generated using the flexible align method within the GALAHAD module, implemented in SYBYL8.1 software. The most significant pharmacophore hypothesis, characterized by the conflicting demands of maximizing pharmacophore consensus, maximizing steric consensus, and minimizing energy, consisted of one positive nitrogen center, one donor atom center, two acceptor atom centers, and two hydrophobic groups. The most active compound in each class training set showed a good fit with all features of the pharmacophore proposed. The resulting models also had something in common with the hypothesis using the Catalyst software reported in other publications. These alpha(1A) pharmacophore models could predict compounds well, both in the training set and the test set. The pharmacophore models were also validated by an external dataset using a portion of the ZINC database. A 3D-QSAR model using the pharmacophore model to align the compounds was established in this study. The CoMFA model with the cross-validated q(2) value of 0.735 revealed that the model was valid. Our research provides a valuable tool for designing new therapeutic compounds with desired biological activity.

Comparative Molecular Field Analysis (CoMFA) and Comparative Molecular Similarity Indices Analysis (CoMSIA) Studies on α1A-Adrenergic Receptor Antagonists Based on Pharmacophore Molecular Alignment

The α1A-adrenergic receptor (α1A-AR) antagonist is useful in treating benign prostatic hyperplasia, lower urinary tract symptoms, and cardiac arrhythmia. Three-dimensional quantitative structure-activity relationship (3D-QSAR) studies were performed on a set of α1A-AR antagonists of N-aryl and N-nitrogen class. Statistically significant models constructed from comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were established based on a training set of 32 ligands using pharmacophore-based molecular alignment. The leave-oneout cross-validation correlation coefficients were q2 CoMFA = 0.840 and q2 CoMSIA = 0.840. The high correlation between the cross-validated/predicted and experimental activities of a test set of 12 ligands revealed that the CoMFA and CoMSIA models were robust (r2 pred/CoMFA = 0.694; r2 pred/CoMSIA = 0.671). The generated models suggested that electrostatic, hydrophobic, and hydrogen bonding interactions play important roles between ligands and receptors in the active site. Our study serves as a guide for further experimental investigations on the synthesis of new compounds. Structural modifications based on the present 3D-QSAR results may lead to the discovery of other α1A-AR antagonists.

Novel naftopidil-related derivatives and their biological effects as alpha1-adrenoceptors antagonists and antiproliferative agents

Eleven novel naftopidil-related compounds that contain amide and indole groups were designed and synthesized. The biological effects of these compounds on three α1-adrenoceptor subtypes and cancerous human prostate cell lines (PC-3, DU-145, and LNCaP) were determined. Compounds 2, 3, 5, 11, and 12 exhibited an α1-adrenoceptor antagonistic activity, whereas compounds 9, 10, and 12 displayed moderate antiproliferative activities. Compound 3 exhibited a significant α(1D/1A) blocking activity in isolated rat tissues (97.7- and 64.6-fold selective for α(1D) and α(1A) compared with α(1B)) but not a relevant cytotoxic activity. Compound 12 demonstrated a potent and selective α(1D/1A) antagonistic activity (47.9- and 19.1-fold for α(1D) and α(1A) compared with α1B) and a potent antiproliferative activity in PC-3 cells (IC50 = 15.70 μM). Further testing confirmed that compound 12 inhibited the growth of PC-3 cells by inducing apoptosis and G0/G1 cell cycle arrest, which was mediated by α1-adrenoceptor. Therefore, compound 12 is a potential multipotent agent that can act as an effective α1-adrenoceptor subtype antagonist for treating benign prostatic hyperplasia and a preventive medication against human prostate cancer.

Metabolic profile of puerarin in rats after intragastric administration of puerarin solid lipid nanoparticles

Puerarin has multiple pharmacological effects and is widely prescribed for patients with cardiovascular diseases including hypertension, cerebral ischemia, myocardial ischemia, diabetes mellitus, and arteriosclerosis. We have successfully prepared puerarin-loaded solid lipid nanoparticles (Pue-SLNs) for oral administration. Pue-SLNs are prepared using monostearin, soya lecithin, and poloxamer 188. SLNs may alter the course of puerarin absorption predominantly to and through lymphatic routes and regions, presumably following a transcellular path of lipid absorption, especially by enterocytes and polar epithelial cells of the intestine. The alteration of absorption might influence the metabolic profile of puerarin when incorporated into SLNs. In the present study, we investigated the metabolic profile of puerarin in rat plasma and urine using rapid resolution liquid chromatography–tandem mass spectrometry after a single-dose intragastric administration of Pue-SLNs in comparison with puerarin suspension. Two glucuronidated metabolites of puerarin, puerarin-4′-O-glucuronide and puerarin-7-O-glucuronide, were detected in rat plasma and urine after intragastric administration of Pue-SLNs, with the latter acting as the major metabolite. Similar results were found in rat plasma and urine after intragastric administration of puerarin suspension. The results suggest that incorporation of puerarin into SLNs does not change either the position of glucuronidation or the metabolic pathway of puerarin in rats.

Determination of naftopidil enantiomers in rat plasma using chiral solid phases and pre-column derivatization high-performance liquid chromatography.

Two bioanalytical HPLC methods (chiral solid phases (CSPs) HPLC and pre-column derivatization HPLC) were developed and validated for the determination of naftopidil enantiomers in rat plasma. Analytes were extracted from biomaterials by liquid-liquid extraction. The pre-column derivatization HPLC method employed (+)-diacetyl-L-tartaric anhydride (DATAN) as the pre-column derivatization reagent, and subsequent separation of diastereomers was conducted on an Agilent Hypersil ODS column with a mixture of methanol-acetonitrile-phosphate buffer (pH 4.1; 20 mM) (40:30:30, v/v/v) flowing at 1 mL/min as the mobile phase. The CSPs HPLC method utilized a Chiralpak IA column with a mobile phase of methanol-acetonitrile-acetate buffer (pH 5.3; 5 mM) (50:25:25, v/v/v) flowing at 0.5 mL/min. In both methods, the analytes were monitored using a fluorescence detector with an excitation wavelength of 290 nm and an emission wavelength of 340 nm. Both methods were consistent (RSD<15% by the derivatization method and<10% by the CSPs method) and linear (r>9950). Compared to the pre-column derivatization method, the CSPs method had lower quantification limits (10.6/9.6 ng/mL of (+)-/(-)-naftopidil by derivatization method and 1.1/1.8 ng/mL of (+)-/(-)-naftopidil by CSPs method), and was simpler to carry out. The validated CSPs method was successfully applied in a pharmacokinetic study of naftopidil enantiomers in rats, which showed that pharmacokinetic parameters of (+)- and (-)-NAF after intravenous administration of (±)-NAF were similar.

Pharmaceutical evaluation of naftopidil enantiomers: Rat functional assays in vitro and estrogen/androgen induced rat benign prostatic hyperplasia model in vivo

Naftopidil (NAF) is a α1D/1A adrenoceptor selective drug used for the treatment of both benign prostatic hyperplasia and lower urinary tract symptoms (BPH/LUTS). However, NAF is used as a racemate in clinic. To compare the differences and similarities among two enantiomers and racemate, pharmacological activities were evaluated through rat functional assays in vitro and estrogen/androgen (E/T) induced rat BPH model in vivo. NAF and the two enantiomers showed similar blocking activity on α1 receptor. S-NAF exhibited more α1D/1A adrenoceptor subtype selectivity than R-NAF and the racemate. The selectivity ratios pA2 (α1D)/pA2 (α1B) and pA2 (α1A)/pA2 (α1B) were 40.7- and 16.2-fold, respectively. NAF and its enantiomers effectively prevented the development of rat prostatic hyperplasia via suppressing the increase of the prostatic wet weight, visually. The quantitative analysis of the relative acinus volume, relative stroma volume, relative epithelial volume, epithelial height and expression of proliferating cell nuclear antigen (PCNA) and α-smooth muscle actin (α-SMA) were carried out. S-NAF showed an advantage on the effect of inhibiting prostate wet weight and stroma volume over R-NAF and racemate NAF (P<0.05). Nevertheless, no other significant difference was observed between these two enantiomers. In conclusion, both R-NAF and S-NAF not only relax prostate muscle but also inhibit the prostate growth, thus relieve BPH.

X-ray Crystallography, DFT Calculations and Molecular Docking of Indole-Arylpiperazine Derivatives as α1A-Adrenoceptor Antagonists.

Indole-arylpiperazine derivatives have exhibited good selectivity for the α1A-adrenoceptor, but the structure-activity-binding mechanism relationship remains unclear. In the current study, three compounds (1, 2 and 3) were investigated through single-crystal X-ray diffraction analysis, density functional theory (DFT) calculations and molecular docking using a homology model of the α1A receptor. Compounds 1 and 3 form H-bonds networks to stabilize their three-dimensional structures, while C–H···π interactions play a significant role in the packing of 2. Based on DFT-optimized conformations, the HOMO-LUMO energy gaps and molecular electrostatic potential (MEP) were theoretically calculated at the B3LYP/6-311G (d, p) level of theory. Chemical reactivity increases in the order of 3 < 2 < 1, and the maximum positive region of the MEP maps is mainly localized over the NH group. The binding mechanisms of ligand-α1A-adrenoceptor complexes were illustrated by molecular docking. Binding to Gln177 of the second extracellular loop region via hydrogen bonds is likely to be essential for α1A-selective antagonists. The present work sheds light on the studies of structure-activity-binding mechanism and aids in the design of α1A antagonists with high selectivity.

Enantiospecific determination of naftopidil by RRLC–MS/MS reveals stereoselective pharmacokinetics and tissue distributions in rats

Naftopidil (NAF) is used as a racemate to treat benign prostatic hyperplasia (BPH) and to prevent prostate cancer. However, racemic NAF has low bioavailability; therefore, it is commonly administered at higher clinical dosages compared to other therapeutic BPH drugs. Differences in interactions between individual enantiomers and biological macromolecules may result in variations in pharmacokinetics and dispositions. This study aimed to investigate the pharmacokinetics, bioavailability and tissue distributions of NAF enantiomers in rats after intragastric administration of the individual enantiomers. A rapid and sensitive liquid chromatography coupled with triple-quadrupole mass spectrometric method (RRLC-MS/MS) was developed and validated for determination of NAF enantiomers in rat plasma, tissues, urine and feces. After intragastric administration, S(-)-NAF in plasma [maximum concentration (C(max)) = 186.4 ng/mL, area under the curve from 0 h to 24h (AUC(0-24 h)) = 877.9 ng h/mL] was significantly higher than that of R(+)-NAF (C(max) = 133.2 ng/mL, AUC(0-24 h) = 602.1 ng h/mL). Moreover, S(-)-NAF bioavailability was twice that of R(+)-NAF. R(+)-NAF distributions in the prostate, liver, and kidney were significantly higher than S(-)-NAF distributions (R/S ratios of 3.16, 1.33, and 2.90, respectively). These data reveal the stereoselective pharmacokinetic profiles of the two enantiomers in rats.