Comprehensive transcriptomics and metabolomics analyses reveal that hyperhomocysteinemia is a high risk factor for coronary artery disease in a chinese obese population aged 40-65: a prospective cross-sectional study.

BACKGROUND: Clinical observations suggest a complex relationship between obesity and coronary artery disease (CAD). This study aimed to characterize the intermediate metabolism phenotypes among obese patients with CAD and without CAD. METHODS: Sixty-two participants who consecutively underwent coronary angiography were enrolled in the discovery cohort. Transcriptional and untargeted metabolomics analyses were carried out to screen for key molecular changes between obese patients with CAD (CAD obese), without CAD (Non-CAD obese), and Non-CAD leans. A targeted GC-MS metabolomics approach was used to further identify differentially expressed metabolites in the validation cohorts. Regression and receiver operator curve analysis were performed to validate the risk model. RESULTS: We found common aberrantly expressed pathways both at the transcriptional and metabolomics levels. These pathways included cysteine and methionine metabolism and arginine and proline metabolism. Untargeted metabolomics revealed that S-adenosylhomocysteine (SAH), 3-hydroxybenzoic acid, 2-hydroxyhippuric acid, nicotinuric acid, and 2-arachidonoyl glycerol were significantly elevated in the CAD obese group compared to the other two groups. In the validation study, targeted cysteine and methionine metabolomics analyses showed that homocysteine (Hcy), SAH, and choline were significantly increased in the CAD obese group compared with the Non-CAD obese group, while betaine, 5-methylpropanedioic acid, S-adenosylmethionine, 4-PA, and vitamin B2 (VB2) showed no significant differences. Multivariate analyses showed that Hcy was an independent predictor of obesity with CAD (hazard ratio 1.7; 95%CI 1.2-2.6). The area under the curve based on the Hcy metabolomic (HCY-Mtb) index was 0.819, and up to 0.877 for the HCY-Mtb.index plus clinical variables. CONCLUSION: This is the first study to propose that obesity with hyperhomocysteinemia is a useful intermediate metabolism phenotype that could be used to identify obese patients at high risk for developing CAD.

Azilsartan ameliorates ventricular hypertrophy in rats suffering from pressure overload-induced cardiac hypertrophy by activating the Keap1-Nrf2 signalling pathway.

OBJECTIVES: Investigate if azilsartan protects against myocardial hypertrophy by upregulating nuclear factor erythroid 2-related factor 2 (Nrf2)-mediated pathways. METHODS: Abdominal aortic constriction (AAC)-induced cardiac hypertrophy in rats was applied. Azilsartan or vehicle was administered daily for 6 weeks in sham or AAC rats. Cardiac morphology and ventricular function were determined. Azilsartan effects upon neonatal rat cardiomyocyte (NRCM) hypertrophy and molecular mechanisms were studied in angiotensin (Ang) II-stimulated NRCMs in vitro. Nrf2-small interfering RNA (siRNA) was used to knockdown Nrf2 expression. Messenger RNA (mRNA)/protein expression of Kelch-like erythroid cell-derived protein (Keap)1 and Nrf2 and its downstream antioxidant enzymes was determined by real-time reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. KEY FINDINGS: Azilsartan treatment ameliorated cardiac hypertrophy/fibrosis significantly in AAC rats. Azilsartan increased expression of Nrf2 protein but decreased expression of Keap1 protein. Upregulation of protein expression of Nrf2's downstream antioxidant enzymes by azilsartan treatment was observed. Azilsartan inhibited Ang II-induced NRCM hypertrophy significantly and similar effects on the Keap1-Nrf2 pathway were observed in vivo. Nrf2 knockdown markedly counteracted the beneficial effects of azilsartan on NRCM hypertrophy and the Keap1-Nrf2 pathway. CONCLUSIONS: Azilsartan restrained pressure overload-induced cardiac remodelling by activating the Keap1-Nrf2 pathway and increasing expression of downstream antioxidant enzymes to alleviate oxidative stress.

Aspirin-omitted dual antithrombotic therapy in non-valvular atrial fibrillation patients presenting with acute coronary syndrome or undergoing percutaneous coronary intervention: results of a meta-analysis.

AIMS: To investigate the effects of aspirin-omitted dual antithrombotic therapy (DAT) on myocardial infarction and stent thrombosis in non-valvular atrial fibrillation (NVAF) patients presenting with acute coronary syndrome (ACS) or undergoing percutaneous coronary intervention (PCI). METHODS AND RESULTS: A systematic review and meta-analysis were performed using PubMed to search for randomized clinical trials comparing DAT with triple antithrombotic therapy (TAT) in this setting. Three trials involving 8845 patients were included (4802 and 4043 patients treated with DAT and TAT, respectively). There were no significant differences in all-cause death and stroke between the aspirin-omitted DAT group and TAT group. Otherwise, the incidence of myocardial infarction was significantly higher with aspirin-omitted DAT vs. TAT [odds ratio (OR): 1.29, 95% confidence interval (CI): 1.02-1.63; P = 0.04; I2 = 0%]. Similarly, the incidence of stent thrombosis increased in patients treated with aspirin-omitted DAT vs. TAT (OR: 1.61, 95% CI: 1.02-2.53; P = 0.04; I2 = 0%). The occurrence of major bleeding and clinically relevant non-major bleeding events, as defined by the International Society on Thrombosis and Haemostasis, was significantly lower with aspirin-omitted DAT vs. TAT (OR: 0.61, 95% CI: 0.48-0.78; P = 0.02; I2 = 76%). Similar results were found according to the International Society on Thrombosis and Haemostasis major bleeding, Thrombolysis in Myocardial Infarction major or minor bleeding, and Thrombolysis in Myocardial Infarction major bleeding scales. CONCLUSION: Aspirin-omitted DAT reduces the occurrence of bleeding episodes, with a higher rate of myocardial infarction and stent thrombosis in NVAF patients presenting with ACS or undergoing PCI.

Puerarin ameliorated pressure overload-induced cardiac hypertrophy in ovariectomized rats through activation of the PPARα/PGC-1 pathway.

Estrogen deficiency induces cardiac dysfunction and increases the risk of cardiovascular disease in postmenopausal women and in those who underwent bilateral oophorectomy. Previous evidence suggests that puerarin, a phytoestrogen, exerts beneficial effects on cardiac function in patients with cardiac hypertrophy. In this study, we investigated whether puerarin could prevent cardiac hypertrophy and remodeling in ovariectomized, aortic-banded rats. Female SD rats subjected to bilateral ovariectomy (OVX) plus abdominal aortic constriction (AAC). The rats were treated with puerarin (50 mg·kg-1 ·d-1, ip) for 8 weeks. Then echocardiography was assessed, and the rats were sacrificed, their heart tissues were extracted and allocated for further experiments. We showed that puerarin administration significantly attenuated cardiac hypertrophy and remodeling in AAC-treated OVX rats, which could be attributed to activation of PPARα/PPARγ coactivator-1 (PGC-1) pathway. Puerarin administration significantly increased the expression of estrogen-related receptor α, nuclear respiratory factor 1, and mitochondrial transcription factor A in hearts. Moreover, puerarin administration regulated the expression of metabolic genes in AAC-treated OVX rats. Hypertrophic changes could be induced in neonatal rat cardiomyocytes (NRCM) in vitro by treatment with angiotensin II (Ang II, 1 µM), which was attenuated by co-treatemnt with puerarin (100 µM). We further showed that puerarin decreased Ang II-induced accumulation of non-esterified fatty acids (NEFAs) and deletion of ATP, attenuated the Ang II-induced dissipation of the mitochondrial membrane potential, and improved the mitochondrial dysfunction in NRCM. Furthermore, addition of PPARα antagonist GW6471 (10 µM) partially abolished the anti-hypertrophic effects and metabolic effects of puerarin in NRCM. In conclusion, puerarin prevents cardiac hypertrophy in AAC-treated OVX rats through activation of PPARα/PGC-1 pathway and regulation of energy metabolism remodeling. This may provide a new approach to prevent the development of heart failure in postmenopausal women.

Contributions of Nrf2 to Puerarin Prevention of Cardiac Hypertrophy and its Metabolic Enzymes Expression in Rats.

Previous evidence has suggested that puerarin may attenuate cardiac hypertrophy; however, the potential mechanisms have not been determined. Moreover, the use of puerarin is limited by severe adverse events, including intravascular hemolysis. This study used a rat model of abdominal aortic constriction (AAC)-induced cardiac hypertrophy to evaluate the potential mechanisms underlying the attenuating efficacy of puerarin on cardiac hypertrophy, as well as the metabolic mechanisms of puerarin involved. We confirmed that puerarin (50 mg/kg per day) significantly attenuated cardiac hypertrophy, upregulated Nrf2, and decreased Keap1 in the myocardium. Moreover, puerarin significantly promoted Nrf2 nuclear accumulation in parallel with the upregulated downstream proteins, including heme oxygenase 1, glutathione transferase P1, and NAD(P)H:quinone oxidoreductase 1. Similar results were obtained in neonatal rat cardiomyocytes (NRCMs) treated with angiotensin II (Ang II; 1 µM) and puerarin (100 µM), whereas the silencing of Nrf2 abolished the antihypertrophic effects of puerarin. The mRNA and protein levels of UGT1A1 and UGT1A9, enzymes for puerarin metabolism, were significantly increased in the liver and heart tissues of AAC rats and Ang II-treated NRCMs. Interestingly, the silencing of Nrf2 attenuated the puerarin-induced upregulation of UGT1A1 and UGT1A9. The results of chromatin immunoprecipitation-quantitative polymerase chain reaction indicated that the binding of Nrf2 to the promoter region of Ugt1a1 or Ugt1a9 was significantly enhanced in puerarin-treated cardiomyocytes. These results suggest that Nrf2 is the key regulator of antihypertrophic effects and upregulation of the metabolic enzymes UGT1A1 and UGT1A9 of puerarin. The autoregulatory circuits between puerarin and Nrf2-induced UGT1A1/1A9 are beneficial to attenuate adverse effects and maintain the pharmacologic effects of puerarin.

Nrf2 Is a Key Regulator on Puerarin Preventing Cardiac Fibrosis and Upregulating Metabolic Enzymes UGT1A1 in Rats.

Puerarin is an isoflavone isolated from Radix puerariae. Emerging evidence shown that puerarin possesses therapeutic benefits that aid in the prevention of cardiovascular diseases. In this study, we evaluated the effects of puerarin on oxidative stress and cardiac fibrosis induced by abdominal aortic banding (AB) and angiotensin II (AngII). We also investigated the mechanisms underlying this phenomenon. The results of histopathological analysis, as well as measurements of collagen expression and cardiac fibroblast proliferation indicated that puerarin administration significantly inhibited cardiac fibrosis induced by AB and AngII. These effects of puerarin may reflect activation of Nrf2/ROS pathway. This hypothesis is supported by observed decreases of reactive oxygen species (ROS), decreases Keap 1, increases Nrf2 expression and nuclear translocation, and decreases of collagen expressions in cardiac fibroblasts treated with a combination of puerarin and AngII. Inhibition of Nrf2 with specific Nrf2 siRNA or Nrf2 inhibitor brusatol attenuated anti-fibrotic and anti-oxidant effects of puerarin. The metabolic effects of puerarin were mediated by Nrf2 through upregulation of UDP-glucuronosyltransferase (UGT) 1A1. The Nrf2 agonist tBHQ upregulated protein expression of UGT1A1 over time in cardiac fibroblasts. Treatment with Nrf2 siRNA or brusatol dramatically decreased UGT1A1 expression in puerarin-treated fibroblasts. The results of chromatin immunoprecipitation-qPCR further confirmed that puerarin significantly increased binding of Nrf2 to the promoter region of Ugt1a1. Western blot analysis showed that puerarin significantly inhibited AngII-induced phosphorylation of p38-MAPK. A specific inhibitor of p38-MAPK, SB203580, decreased collagen expression, and ROS generation induced by AngII in cardiac fibroblast. Together, these results suggest that puerarin prevents cardiac fibrosis via activation of Nrf2 and inactivation of p38-MAPK. Nrf2 is the key regulator of anti-fibrotic effects and upregulates metabolic enzymes UGT1A1. Autoregulatory circuits between puerarin and Nrf2-regulated UGT1A1 attenuates side effects associated with treatment, but it does not weaken puerarin's pharmacological effects.

Puerarin-7-O-glucuronide, a water-soluble puerarin metabolite, prevents angiotensin II-induced cardiomyocyte hypertrophy by reducing oxidative stress.

This study aimed to investigate the anti-oxidant and anti-hypertrophic effects of puerarin-7-O-glucuronide, a water-soluble puerarin metabolite. The anti-oxidant effects of puerarin-7-O-glucuronide were assessed by measurement of intracellular superoxide levels, total superoxide dismutase (SOD) activity, total anti-oxidant capacity, and glutathione (GSH)/glutathione disulfide (GSSG) ratio in cultured neonatal rat cardiomyocytes (NRCMs) stimulated with the xanthine oxidase (XO)/xanthine (X) system or angiotensin II. The activity of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and expression of NADPH oxidase subunits p22phox and p47phox were determined. The anti-hypertrophic effects of puerarin-7-O-glucuronide in angiotensin II-challenged NRCMs were characterized by changes in cell morphology and expression of hypertrophic genes. In the pharmacokinetic study, the plasma concentration of puerarin-7-O-glucuronide was determined by rapid resolution-liquid chromatography-tandem mass spectrometry (RR-LC-MS/MS). Puerarin-7-O-glucuronide prevented XO/X-induced increase in intracellular superoxide production and decreases in total SOD activity, GSH/GSSG ratio, and total anti-oxidant capacity. Puerarin-7-O-glucuronide also reversed angiotensin II-induced increases in intracellular superoxide production and NADPH oxidase activity and decreases in total SOD activity. These anti-oxidant effects of puerarin-7-O-glucuronide were accompanied by downregulation of p22phox and p47phox. Furthermore, puerarin-7-O-glucuronide prevented angiotensin II-induced increases in cell surface area and perimeter, as well as changes in Nppa, Myh7, and Myh6. In the pharmacokinetic study, puerarin-7-O-glucuronide was cleared with a half-life of 0.94 h after intravenous administration. Puerarin could be detected in rat plasma, albeit in low concentration, as early as 5 min after intravenous administration of puerarin-7-O-glucuronide. These anti-oxidant and anti-hypertrophic properties of puerarin-7-O-glucuronide were similar to those of its parent compound puerarin. These results support the development of puerarin-7-O-glucuronide as a novel pharmaceutical agent for therapeutic application.

Human UDP-Glucuronosyltransferase 2B4 and 2B7 Are Responsible for Naftopidil Glucuronidation in Vitro.

Naftopidil (NAF) is widely used for the treatment of benign prostatic hyperplasia and prevention of prostate cancer in elderly men. These patients receive a combination of drugs, which involves high risk for drug-drug interaction. NAF exhibits superior efficacy but must be administered at a much higher dosage than other therapeutic drugs. We previously showed that extensive glucuronidation of NAF enantiomers caused poor bioavailability. However, the metabolic pathway and mechanism of action of NAF enantiomer remain to be elucidated. The present study was performed to identify the human UDP-glucuronosyltransferases (UGTs) responsible for the glucuronidation of NAF enantiomers and to investigate the potential inhibition of UGT activity by NAF. The major metabolic sites examined were liver and kidney, which were compared with intestine. Screening of 12 recombinant UGTs showed that UGT2B7 primarily contributed to the metabolism of both enantiomers. Moreover, enzyme kinetics for R(+)-NAF, UGT2B7 (mean Km, 21 µM; mean Vmax, 1043 pmol/min/mg) showed significantly higher activity than observed for UGT2B4 and UGT1A9. UGT2B4 (mean Km, 55 µM; mean Vmax, 1976 pmol/min/mg) and UGT2B7 (mean Km, 38 µM; mean Vmax, 1331 pmol/min/mg) showed significantly higher catalysis of glucuronidation of S(-)-NAF than UGT1A9. In human liver microsomes, R(+)-NAF and S(-)-NAF also inhibited UGT1A9: mean Ki values for R(+)-NAF and S(-)-NAF were 10.0 µM and 11.5 µM, respectively. These data indicate that UGT2B7 was the principal enzyme mediating glucuronidation of R(+)-NAF and S(-)-NAF. UGT2B4 plays the key role in the stereoselective metabolism of NAF enantiomers. R(+)-NAF and S(-)-NAF may inhibit UGT1A9. Understanding the metabolism of NAF enantiomers, especially their interactions with metabolic enzymes, will help to elucidate potential drug-drug interactions and to optimize the administration of this medicine.

Impact of Timing following Acute Myocardial Infarction on Efficacy and Safety of Bone Marrow Stem Cells Therapy: A Network Meta-Analysis.

Background. The optimal timing for Bone Marrow Stem Cells (BMCs) therapy following acute myocardial infarction (AMI) remains unclear. Aims. To synthesize the evidence from trials using a multiple-treatment comparison method, thereby permitting a broader comparison across multiple timing of BMCs therapy. Methods and Results. Randomized controlled trials in patients with AMI receiving BMCs therapy were identified from PubMed, Ovid LWW, BIOSIS Previews, and the Cochrane Library through January 2015. 2 035 patients of 31 studies included in our analysis were allocated to 5 groups' treatments: 1~3 days, 4~7 days, 8~14 days, 15~30 days, or placebo/control group. The multiple-treatment meta-analysis showed that 4~7 days' group could lead to significantly increased left ventricular ejection fraction (LVEF) as compared with control (mean of MDs and 95% CI: 6 months, 3.05 (0.92~5.25); 12 months, 4.18 (2.30~5.84)). Only 4~7 days led to significant reduction of MACEs compared with control (OR and 95% CI 0.34 (0.13~0.96)) for 12-months follow-up. In simulated comparisons, the 4~7 days' group ranked better than other timing groups for improvement of LVEF or reduction of the incidence of major adverse cardiac events. Conclusions. 4~7 days after AMI might be the optimal timing for cell therapy in terms of efficacy or safety.

Effectiveness and safety of selected bone marrow stem cells on left ventricular function in patients with acute myocardial infarction: a meta-analysis of randomized controlled trials.

BACKGROUND: Concerns regarding the use of selected bone marrow stem cells (BMSCs) in the field of cardiac repair after acute ischemic events have been raised. The current meta-analysis aimed to assess the efficacy and safety of selected BMSC transplantation in patients with acute myocardial infarction (AMI) based on published randomized controlled trials (RCTs). METHODS: A systematic literature search of PubMed, Ovid LWW, BIOSIS Previews, and the Cochrane library from 1990 to 2014 was conducted. Results from RCTs involving subjects with AMI receiving selected BMSC therapy and followed up for at least 6 months were pooled. RESULTS: Eight trials with a total of 262 participants were included. Data were analyzed using a random effects model. Overall, selected BMSC therapy improved left ventricular ejection fraction (LVEF) by 3.17% (95% confidence interval [CI] 0.57-5.76, P=0.02), compared with the controls. There were trends toward reduced left ventricular end-systolic volume (LVESV) and fewer major adverse cardiac events (MACEs). Subgroup analysis revealed a significant difference in LVEF in favor of selected BMSC therapy with bone marrow mesenchymal stem cells (BMMSCs) as the cell type. CONCLUSIONS: Transplantation of selected BMSCs for patients with AMI is safe and induces a significant increase in LVEF with a limited impact on left ventricular remodeling.

MicroRNA-26 was decreased in rat cardiac hypertrophy model and may be a promising therapeutic target.

MicroRNA (miR)-26 was found to be downregulated in cardiac diseases. In this study, the critical role of miR-26 in myocardial hypertrophy in both in vivo and in vitro was investigated. Sixteen male Wistar rats that underwent sham or transverse abdominal aortic constriction (TAAC) surgery were divided into control or TAAC group. Cardiomyocytes were isolated from neonatal Sprague-Dawley rats. Our study demonstrated that miR-26a/b was downregulated in both TAAC rat model and cardiomyocytes. The results of luciferase assays also suggested that glycogen synthase kinase 3ß (GSK3ß) may be a direct target of miR-26. The overexpression of miR-26 attenuated GSK3ß expression and inhibited myocardial hypertrophy. The downregulation of miR-26 reversed these effects. Furthermore, silence of GSK3ß gene phenocopied the anti-hypertrophy effects of miR-26, whereas overexpression of this protein attenuated the effects of miR-26. Taken together, these data suggest that miR-26 regulates pathological structural changes in the rat heart, which may be associated with suppression of the GSK3ß signaling pathway, and implicate the potential application of miR-26 in diagnosis and therapy of cardiac hypertrophy.

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.

UDP-glucuronosyltransferase 1A1 is the principal enzyme responsible for puerarin metabolism in human liver microsomes.

Puerarin has multiple pharmacological effects and is widely prescribed for patients with cardiovascular diseases, including hypertension, cerebral ischemia, myocardial ischemia, diabetes mellitus, and arteriosclerosis. While puerarin is a useful therapeutic agent, its mechanisms of action have not been well defined. Understanding puerarin metabolism, in particular its interactions with metabolizing enzymes, will contribute to our understanding of its toxic and therapeutic effects and may help to elucidate potential negative drug-drug interactions. In this study, the major metabolite of puerarin was obtained from the urine of rats administered puerarin, by a semi-preparative high-performance liquid chromatography method. The major metabolite was identified as puerarin-7-O-glucuronide. In vitro, we used a UDP-glucuronosyltransferase (UGT) reaction screening method with 12 recombinant human UGTs to demonstrate that formation of puerarin-7-O-glucuronide was catalyzed by UGT1A1, 1A9, 1A10, 1A3, 1A6, 1A7, and 1A8. UGT1A1, 1A9, and 1A10 significantly catalyzed puerarin-7-O-glucuronide formation, and the activity of UGT1A1 was significantly higher than those of 1A9 and 1A10. The V (max) of UGT1A1 was two- to threefold higher than the levels of UGT1A9 or 1A10, with a lower K ( m ) value and a higher V (max)/K ( m ) value. The kinetics of puerarin-7-O-glucuronide formation catalyzed by UGT1A1 were similar to those of the pooled human liver microsomes (HLMs), with V (max) values of 186.3 and 149.2 pmol/min/mg protein, and K ( m ) values of 811.3 and 838.9 µM, respectively. Furthermore, bilirubin and ß-estradiol, probe substrates for UGT1A1, significantly inhibited the formation of puerarin-7-O-glucuronide in HLMs.

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).

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.

[Molecular mechanism of protective effect of puerarin solid lipid nanoparticle on cerebral ischemia-reperfusion injury in gerbils].

OBJECTIVE: To investigate the effects of puerarin solid lipid nanoparticle on fore brain ischemic-reperfusion injury in gerbils and it's mechanisms. METHODS: Gerbils were randomly divided into 4 groups: sham group, cerebral ischemia-reperfusion injury group, puerarin solid lipid nanoparticle group and puerarin injection control group. The gerbils' cerebral ischemia-reperfusion injury model was constructed with ligating bilater carotids method. The histomorphology and Bcl-2, Caspase-3 and HSP70 expressions were detected by HE dyeing and immunohistochemical method. RESULTS: After 24 h ischemia and reperfusion in gerbils, the level of Bcl-2 and HSP70 expressions in puerarin solid lipid nanoparticle group increased (P < 0.01) compared with the ischemic-reperfusion model group, and the level of Caspase-3 expression decreased (P < 0.01). The same results was consistent in puerarin injection control group. CONCLUSIONS: Puerarin solid lipid nanoparticle group can protect the cerebral ischemia-reperfusion injury in gerbils, which may be related to the upregulation of Bcl-2 and HSP70 expression and downregulation of Caspase-3 expression.

Metabolites of puerarin identified by liquid chromatography tandem mass spectrometry: similar metabolic profiles in liver and intestine of rats.

Puerarin is a major active ingredient of Pueraria radix. Puerarin may exert its medicinal functions in part via its metabolites. In this study, we identified these metabolites to better understand and elucidate puerarin's metabolic pathway. Puerarin was intravenously administered to rats and then metabolites in plasma samples were identified by rapid resolution liquid chromatography electrospray ionization-collision induced dissociation tandem mass spectrometry (RRLC-ESI-CID-MS/MS). Chromatography was conducted on a Zorbax SB C18 column (2.1x100 mm, 1.8 microm) at 30 degrees C, with a gradient mobile phase consisting of 0.05% formic acid and acetonitrile, a flow rate of 0.2 mL min(-1), and a total run time of 14 min. MS/MS acquisition parameters were as follows: positive ionization mode, dry gas: nitrogen, 10 L min(-1), dry temperature: 350 degrees C, nebulizer: 40 psi, capillary: -3500 V, scan range: 250-800. The autoMS, manual, or multiple reaction monitoring mode was selected as required. Two glucuronidated metabolites of puerarin (M1 and M2) were detected. M1 and M2 are presumed to be puerarin-7-O-glucuronide and puerarin-4'-O-glucuronide, respectively, and M2 likely is suspected to be the major metabolite because it represented the predominate peak. Kinetic studies of metabolites demonstrated that M1 and M2 were detected in rat plasma at 5 min after intravenous administration of puerarin, the levels of M1 and M2 then reached their peaks at 10-15 and 15-30 min, respectively. The metabolic profiles were similar in rat liver and intestine investigated by in situ liver and intestine perfusion, indicating that no metabolic regioselectivity of puerarin occurs in the two organs.