Puerarin Concentration Research Articles

Miniature and cost‐effective self‐powered triboelectric sensing system toward rapid detection of puerarin concentration

AbstractThe detection of puerarin concentration is an essential capability to study the functional role of the Pueraria root as a natural medicine and dietary source in the treatment of cardiovascular diseases and liver protection. Current methods to detect and measure puerarin concentration, such as ultraviolet–visible spectrophotometry (UV), are bulky, require an external power supply, and are inconvenient to use. Here, we propose a triboelectric puerarin‐detecting sensor (TPDS) which is based on liquid–solid contact electrification, in which liquid–solid interactions generate rapid electrical signals in only 0.4 ms to enable real‐time detection of puerarin concentration in water droplets. The electrical signal of the TPDS decreases with an increase of puerarin concentration, and the sensitivity of the approach is 520 V·(μg/mL)−1. The TPDS represents a miniature and cost‐effective sensor that is 0.2% of the size and 0.01% of the cost of a UV spectrophotometer. Our theoretical analysis verified that the puerarin concentration in droplets can effectively regulate the electronic structure, where higher concentrations of puerarin lead to a narrower energy bandgap, which allows the TPDS to detect puerarin concentration without the need for an external power supply. The TPDS therefore provides a route for the development of a portable and self‐powered method to measure the concentration of an active ingredient in droplets through the conversion of natural energy.image

Changes in the content of Puerarin-PLGA nanoparticles in mice under the influence of alcohol and analysis of their antialcoholism.

To observe the metabolic changes and antialcoholic effect of Puerarin-PLGA nanoparticles (PUE-NP) in mice. PUE-NP was prepared and characterized by particle size distribution and morphology. The mouse models with acute alcoholism were established to observe their behavioral changes after alcohol poisoning. The expressions of biologically active enzymes such as CRE, BUN, AST, ALT in serum and SOD and TLR4 in liver of mice in each group were detected, and the pathological changes in liver and kidney tissues were observed by HE staining. The PUE-NP metabolism in mice was determined by in vitro release assay and HPLC. PUE-NP nanoparticles had good morphology and structure, and the mouse models with alcohol poisoning were established successfully. Compared with alcohol group, puerarin and PUE-NP increased the disappearance latency time of righting reflex, and the recovery time of righting reflex was significantly shortened. Water maze results showed that Puerarin and PUE-NP had inhibitory effect on impaired memory. HPLC results showed that PUE-NP reached its peak in mice after 1 h, and the content percentage was twice that of puerarin preparation alone, and the distribution time of puerarin concentration in vivo was prolonged, indicating that PLGA nanoparticles had a loading and slow-release effect on puerarin and increased the bioavailability of puerarin in mice. In addition, compared with the alcohol group, Puerarin and PUE-NP improved serum ALT, AST, CRE, and BUN levels in mice, enhanced SOD activity in liver, and inhibited TLR4 expression. The effect was better in the PUE-NP group than in the Puerarin group. PUE-NP delayed the release and metabolism of Puerarin and had better effect in the treatment of the alcoholic liver and kidney injury.

Inhibitory role of puerarin on the A549 lung cancer cell line.

Although more and more drugs had been proved to be effective in controlling tumor cells, lung cancer was still the leading cause of cancer-related deaths all over the world. This study aimed to investigate the effect and mechanism of puerarin on the invasion and metastasis of A549 lung cancer cell line. A medium containing puerarin was prepared according to the gradient concentration, and 10, 20, and 40 µmol/L were selected as the experimental group (low, medium, and high concentration groups, respectively) according to the cytotoxicity experiment. Meanwhile, 0 µmol/L was used as the control group. Following administration, metastasis-related indexes were detected by the cell scratch test, cell migration test, gene difference detection, and western blotting. 24 hours after administration, the cell scratch and Transwell showed that the migration ability of A549 cells decreased with the increasing puerarin concentration. The polymerase chain reaction (PCR) and western blotting results demonstrated that the expression of the cell invasion and metastasis-related factor, matrix metallopeptidase 9 (MMP9), was negatively correlated with drug concentration. Further investigation demonstrated that the phosphorylation of extracellular signal-regulated kinase (ERK) was also inhibited. Puerarin can inhibit the expression of invasion and metastasis-related factors by inhibiting the phosphorylation of ERK.

MiR-34a-5p/Sirt1 axis: A novel pathway for puerarin-mediated hepatoprotection against benzo(a)pyrene

Benzo[a]pyrene (BaP) as a carcinogen induces oxidative stress and inflammation, causing health problems including liver damage. Puerarin (a natural flavonoid) is traditionally used to provide hepatoprotective effects. This research was established to meet the rising demand for effective therapies/treatments against hepatic diseases and investigate the mechanism underlying the protective actions of puerarin against BaP-induced liver damage. In mice, puerarin combated effectively the detrimental changes in liver weight, color and function indices caused by BaP. In HepG2 cells, puerarin alleviated BaP-induced cell death, oxidative stress and inflammation, and such effects were positively correlated with puerarin's concentration (12.5–50 μM). Mechanistic studies revealed that BaP induced low Sirt1 expression and high miR-34a-5p expression, and puerarin treatment alleviated these changes. Oxidative stress and inflammation induced by BaP were almost eliminated when miR-34a-5p was silenced. Inhibiting miR-34a-5p or overexpressing Sirt1 had a similar effect to puerain treatment. Overexpression of miR-34a-5p and inhibition of Sirt1 reduced the protective effect of puerarin. Collectively, miR-34a-5p participates in the regulation of puerarin's protective function against BaP-induced injury through targeting Sirt1. There is a novel pathway for suppressing oxidative stress and inflammation via miR-34a-5p/Sirt1 axis in puerarin-mediated hepatoprotection, which opens up a new avenue for alternative therapies.

Puerarin Inhibits the Proliferation,Invasion,and Migration of Non-small Cell Lung Cancer Cells through Regulating miR-490/Denticleless E3 Ubiquitin Protein Ligase

Objective To explore the mechanism of puerarin inhibiting the proliferation,invasion,and migration of non-small cell lung cancer cells. Methods A549 cells were cultured and treated with different concentrations of puerarin.The inhibition rate (IR) on cell proliferation was detected by CCK-8,and qRT-PCR was performed to detect the mRNA levels of miR-490 and denticleless E3 ubiquitin protein ligase(DTL).Double luciferase reporter assay was employed to identify the targets of miR-490 and DTL based on the establishment of NC mimic group,miR-490 mimic group,NC inhibitor group,and miR-490 inhibitor group.The cells treated by 20 μmol/L puerarin were classified into six groups:DMSO,puerarin,puerarin+NC inhibitor,puerarin+miR-490 inhibitor,puerarin+miR-490 inhibitor+Si-NC,and puerarin+miR-490 inhibitor+Si-DTL.Transwell was used to detect cell migration and invasion.Western blotting was performed to detect the protein levels of epithelial-mesenchymal transition-related markers E-cadherin,N-cadherin,and Vimentin. Results With the increase in puerarin concentration,the IR gradually elevated (F=105.375,P<0.001),miR-490 expression gradually increased (F=32.919,P<0.001),and DTL expression gradually decreased (F=116.120,P<0.001).Compared with NC mimic group,miR-490 mimic group had decreased luciferase activity (t=7.762,P=0.016),raised miR-490 mRNA level (t=13.319,P<0.001),and declined DTL mRNA level (t=7.415,P=0.002).Compared with those in NC inhibitor group,miR-490 demonstrated decreased mRNA level (t=9.523,P=0.001) and DTL presented increased mRNA level (t=11.305,P<0.001) in miR-490 inhibitor group.Western blotting showed that the protein level of DTL was higher in NC mimic group (t=7.953,P=0.001) than in miR-490 mimic group and higher in miR-490 inhibitor group than in NC inhibitor group (t=10.552,P<0.001).Compared with DMSO group,puerarin group showed up-regulated mRNA level of miR-490 (t=10.255,P=0.001) while down-regulated mRNA level of DTL (t=6.682,P=0.003).Compared with those in puerarin+NC inhibitor group,the mRNA level of miR-490 declined (t=10.995,P<0.001) while that of DTL raised (t=12.478,P<0.001) in puerarin+miR-490 inhibitor group.The mRNA level of miR-490 had no significant difference between puerarin+miR-490 inhibitor+Si-NC group and puerarin+miR-490 inhibitor+Si-DTL group (t=1.081,P=0.341),and that of DTL was lower in the latter group (t=14.321,P<0.001).The protein level of DTL was higher in puerarin+miR-490 inhibitor group than in puerarin+NC inhibitor group (t=11.423,P<0.001),and lower in puerarin+miR-490 inhibitor+Si-DTL group than in puerarin+miR-490 inhibitor+Si-NC group (t=12.080,P<0.001).Compared with DMSO group,puerarin group showed inhibited cell proliferation (F=129.27,P<0.001).The activity of cell proliferation was higher in puerarin+miR-490 inhibitor group than in puerarin+NC inhibitor group (F=75.12,P<0.001),and higher in puerarin+miR-490 inhibitor+Si-NC group than in puerarin+miR-490 inhibitor+Si-DTL group (F=52.59,P<0.001).Compared with DMSO group,puerarin group had suppressed cell migration (t=8.963,P=0.001).The cell migration ability was higher in puerarin+miR-490 inhibitor group than in puerarin+NC inhibitor group (t=12.117,P<0.001) and higher in puerarin+miR-490 inhibitor+Si-NC group than in puerarin+miR-490 inhibitor+Si-DTL group (t=12.934,P<0.001).Puerarin group showed weakened cell invasion ability compared with DMSO group (t=4.710,P=0.009).The cell invasion ability was higher in puerarin+miR-490 inhibitor group than in puerarin+NC inhibitor group (t=13.264,P<0.001) and lower in puerarin+miR-490 inhibitor+Si-DTL group than in puerarin+miR-490 inhibitor+Si-NC group (t=13.476,P<0.001).Compared with DMSO group,puerarin group showed up-regulated protein level of E-cadherin (t=7.137,P=0.002) while down-regulated protein levels of N-cadherin (t=8.828,P=0.001) and vimentin (t=6.594,P=0.003).Compared with those in puerarin+NC inhibitor group,the protein level of E-cadherin (t=12.376,P<0.001) decreased while those of N-cadherin (t=13.436,P<0.001) and vimentin (t=11.467,P<0.001) increased in puerarin+miR-490 inhibitor group.Compared with puerarin+miR-490 inhibitor+Si-NC group,puerarin+miR-490 inhibitor+Si-DTL group up-regulated the protein level of E-cadherin (t=13.081,P<0.001) while down-regulated the protein levels of N-cadherin (t=10.835,P<0.001) and vimentin (t=11.862,P<0.001). Conclusion Puerarin could inhibit the proliferation,invasion,and migration of non-small cell lung cancer cells by up-regulating miR-490 and down-regulating DTL.

Effect of puerarin on osteogenic differentiation of human periodontal ligament stem cells.

ObjectiveTo investigate the effects of the flavonoid, puerarin, on osteogenic differentiation of human periodontal ligament stem cells (PDLSCs).MethodsHuman PDLSCs were isolated from patients undergoing orthodontic treatment, and the cell surface markers CD146, CD34, CD45, and STRO-1 were identified by immunofluorescence. Cell proliferation was detected by MTT assay; alkaline phosphatase (ALP) activity was measured, and calcium deposition was detected by alizarin red staining. PCR was then used to detect the distributions of COL-I, OPN, Runx2, and OCN, genes related to osteogenic differentiation.ResultsStaining was positive for cytokines CD146, CD34, CD45, and STRO-1 in the experimental group; staining was also positive for silk protein, but negative for keratin. After 7 days of culture, exposure to puerarin significantly promoted the level of intracellular ALP; increased puerarin concentration led to increased intracellular ALP. Red mineralized nodules appeared upon exposure to puerarin and the number of nodules was concentration-dependent. PCR analysis revealed that COL-I, OPN, Runx2, and OCN expression levels increased as puerarin concentration increased.ConclusionsExposure to puerarin can promote proliferation and ALP activity in human PDLSCs, thus promoting both molecular and osteogenic differentiation; these findings may provide a theoretical basis for the clinical treatment of periodontal disease with puerarin.

Puerarin Exerts Protective Effects on Wear Particle-Induced Inflammatory Osteolysis.

Wear particle-stimulated inflammatory bone destruction and the consequent aseptic loosening remain major postoperative problems for artificial joints. Studies have indicated that puerarin promotes osteogenesis and alleviates lipopolysaccharide-induced osteoclastogenesis in vitro. However, the underlying molecular mechanism by which puerarin interacts with receptor activator of nuclear factor kappa-B ligand (RANKL)-mediated osteoclast formation in vitro and wear particle-stimulated osteolysis in vivo has not been reported. In this work, the protective effects exerted by puerarin on titanium particle-stimulated bone destruction in vivo and on RANKL-induced osteoclast activation in osteoclastic precursor cells in vitro were investigated. As expected, puerarin significantly inhibited wear particle-mediated bone resorption and proinflammatory cytokine productions in a calvarial resorption model. Additionally, puerarin inhibited RANKL-induced osteoclast activation, bone resorption ability, and F-actin ring formation in vitro as puerarin concentration increased. Furthermore, mechanistic investigation indicated that reduced RANKL-stimulated MEK/ERK/NFATc1 signaling cascades might regulate the protective effect of puerarin. Conclusively, these results indicate that puerarin, a type of polyphenol, might serve as a protective agent to prevent osteoclast-related osteolytic diseases.

Distribution kinetics of puerarin in rat hippocampus after acute local cerebral ischemia

Puerarin, isolated from the roots of Pueraria lobata, is widely used for treating cerebral ischemia in China. The time- and dose-dependent distribution characteristics of puerarin in the ischemic hippocampus are unknown. In this study, puerarin concentration was determined by an indirect competitive ELISA using anti-puerarin monoclonal antibody. Area under the curve (AUC0-120 min) of puerarin (80 mg/kg) in the embolic hippocampus was higher than that in the normal hippocampus; the increase was significant only at 40 and 20 mg/kg. The maximum concentration (Cmax) of puerarin in the embolic hippocampus was higher than that in the normal hippocampus at all doses. The increase in both AUC0-120 min and Cmax was dose-dependent. Time to reach the maximum concentration (Tmax) of puerarin in the embolic and normal hippocampus was similar. Although the mean residence time in the embolic hippocampus differed from that in the normal hippocampus at 40 and 80 mg/kg, it was higher in the embolic hippocampus than in the normal hippocampus at 20 mg/kg. This is the first study to report that the time- and dose-dependent distribution characteristics of puerarin in the normal and embolic hippocampus after middle cerebral artery occlusion in rats dictate puerarin dose and duration to treat stroke.

Experimental factors affecting in vitro recovery of puerarin in microdialysis

: To analyze experimental factors affecting in vitro recovery of puerarin in microdialysis. : Puerarin concentration in microdialysate samples was determined by high performance liquid chromatography. The methods of direct dialysis, retrodialysis and the zero-net flux were used to calculate in vitro recovery, respectively. The effects of perfusate composition, the analyte concentration, perfusate flow rate, medium temperature and stir rates of the dialysis medium on recovery were investigated. : There were significant differences in the recovery values among direct dialysis, retrodialysis and zero-net flux methods. The recovery for 0.9% NaCl solution, Ringer's solution, PBS and anticoagulant dextrose solution as perfusate fluid were (71.25±2.36)%,(73.48±1.41)%,(68.50±2.43)% and (74.98±1.16)%, respectively. The composition of perfusate fluid had significant influence on the recovery(P<0.01). At the same flow rate, recovery was independent of the analyte concentration. At the same concentration, the recovery was decrease with the increasing flow rate in an exponential relationship. The recovery increased with the raising temperature and stir rate of the dialysis medium, and the recovery remained stable when the stir rate reached above 200 rpm. : A study method for in vitro recovery of puerarin in microdialysis has been established, and the recovery of puerarin is affected by calculating methods, perfusate fluids, flow rate, medium temperature and stir rate, but not affected by analyte concentrations.

Effect of puerarin in Longmaining formula on pharmacokinetics-pharmacodynamics correlation in rats with myocardial ischemia

To study the pharmacokinetics of puerarin in compound Longmaining(FFLMN) in normal rats and myocardial ischemia rats, and investigate its correlation with anti-myocardial ischemia effect of FFLMN. Models of myocardial ischemia rats were produced by subcutaneous injection of isoproterenol(ISO), then FFLMN extract solution was administered by gavage. Orbital sinus blood sampling was collected at different time points after gavage. HPLC-UV method was applied to determine the concentration of puerarin in plasma, and compare the difference in pharmacokinetics between normal rats and model rats after application of same dose of FFLMN. Meanwhile, microplate reader was used to determine IL-6 and SOD activities in plasma of different time points, and draw dose-effect curve. The results indicated that the pharmacokinetics of puerarin conformed to the two-compartment model in both normal group and myocardial ischemia model group. In the comparison of main pharmacokinetic parameters between two groups: AUC0-∞=(11.451±3.228) mg•h•L⁻¹,AUC0-t=(14.047±3.765) mg•h•L⁻¹, Cmax=(5.623±1.40) mg•L⁻¹ in normal group; AUC0-∞=(68.849±50.396 9) mg•h•L⁻¹, AUC0-t= (58.312±45.802) mg•h•L⁻¹,Cmax=(18.456±7.517) mg•L⁻¹ in treatment group. The SOD level was significantly increased and IL-6 concentration was significantly decreased in plasma, indicating that as compared with the normal group, puerarin in FFLMN had a higher plasma concentration, slower elimination rate and higher bioavailability. Therefore, puerarin concentration in plasma has correlation with the anti-myocardial ischemia effect of FFLMN, which could increase SOD level and inhibit the release of IL-6.

Effect of puerarin on human choriocarcinoma cells.

ObjectiveTo discuss the effect of puerarin on human choriocarcinoma cells.MethodsSurvival rates under puerarin monotherapy, fluorouracil (5-FU) monotherapy and puerarin in combination with 5-FU were detected by MTT assay. Apoptotic morphology was observed with Hoechst 33258 staining. Apoptosis rates were detected with flow cytometry. Expressions of AKT, mechanistic target of rapamycin (mTOR), and P70S6K mRNAs and phosphorylated proteins were detected by RT-PCR and Western blot. Tumor-bearing mice were administered puerarin and puerarin+5-FU, and serum levels of β-human chorionic gonadotropin (β-HCG) were measured.ResultsProliferation inhibition and apoptosis rates of JEG-3 cells were positively correlated with puerarin concentration, which increased in the puerarin+5-FU group. Expression levels of AKT, mTOR, P70S6K mRNAs, and phosphorylated proteins decreased significantly after action of puerarin at different concentrations. With increasing puerarin concentration, expression of cleaved-caspase-3 in JEG-3 cells increased, whereas that of Bcl-2 decreased. Puerarin significantly inhibited tumor growth in choriocarcinoma-bearing SCID mice. Serum β-HCG levels were significantly lower than those of control group after administration. Magnitude of β-HCG decline was positively correlated with concentration..ConclusionPuerarin+5-FU inhibited proliferation of JEG-3 choriocarcinoma cells and promoted their apoptosis, being associated with the mTOR signaling pathway.

Puerarin exhibits greater distribution and longer retention time in neurons than astrocytes in a co-cultured system.

The phytoestrogen puerarin has been shown to protect neurons and astrocytes in the brain, and is therefore an attractive drug in the treatment of Alzheimer's disease, Parkinson's disease and cerebral ischemia. Whether puerarin exhibits the same biological processes in neurons and astrocytes in vitro has rarely been reported. In this study, cortical neurons and astrocytes of newborn Sprague-Dawley rats were separated, identified and co-cultured in a system based on Transwell membranes. The retention time and distribution of puerarin in each cell type was detected by fluorescence spectrophotometry and fluorescence microscope. The concentration of puerarin in both co-cultured and separately cultured neurons was greater than that of astrocytes. Puerarin concentration reached a maximum 20 minutes after it was added. At 60 minutes after its addition, a scant amount of drug was detected in astrocytes; however in both separately cultured and co-cultured neurons, the concentration of puerarin achieved a stable level of about 12.8 ng/mL. The results indicate that puerarin had a higher concentration and longer retention time in neurons than that observed in astrocytes.

Comparison of two approaches of intestinal absorption by puerarin

IntroductionEverted gut sac (EGS) and in situ single-pass intestinal perfusion (SPIP) have been widely used in the study of drug absorption and biopharmaceutical classification systems (BCS). Furthermore, they could also be applied in the research of drug intestinal first-pass metabolism. Since most of Chinese herbal medicines (CHMs) are orally administrated, it is necessary to study the permeability of active ingredients of CHMs. Thus, we attempted to apply the EGS and SPIP models to study the permeability of puerarin, one of the active marker compounds (AMCs) of Puerariae Radix. MethodsIn the present study, three rat models of ex vivo and in situ, EGS, SPIP, and in situ intestinal perfusion with venous sampling (IPVS), were established to determine the permeability coefficient of puerarin. The apparent permeability coefficient (Papp) was obtained by EGS. The SPIP model was used to determine the effective permeability coefficient (Peff) in different intestinal segments. The blood permeability coefficient (Pblood) was determined by IPVS. Puerarin concentration of perfusion and blood samples were measured by HPLC. ResultsPuerarin could filter into EGS incubated in aqueous extract of Puerariae Radix or puerarin solution. In the SPIP experiment, the concentration effect on Peff was observed in the ileum, but not in the other three intestinal segments. The Pblood was 0.068±0.002×10−5cm/s and was 16-fold lower than the Peff (1.114±0.153×10−5cm/s) in the IPVS experiment at 80μg/mL puerarin. As expected, the Peff (1.24±0.11×10−5cm/s) in SPIP did not differ from the Peff in IPVS. The Papp was 0.199×10−5cm/s at 1200μg/mL puerarin, 10-fold lower than Peff (2.047±0.116×10−5cm/s) in SPIP. DiscussionThree models for permeability were successfully practiced in the study of puerarin absorption and our research strategy will be useful for herbal constituent absorption in the future.

Puerarin Protects AgainstStaphylococcus aureus-Induced Injury of Human Alveolar Epithelial A549 Cells via Downregulating Alpha-Hemolysin Secretion

Alpha-hemolysin, a secreted pore-forming toxin, plays an indispensable role in the pathogenicity of Staphylococcus aureus. In this study, the antimicrobial activity of puerarin against S. aureus was investigated; as a result, puerarin showed no influence on the growth of this organism. However, hemolysis and western blotting assays showed that puerarin concentration dependently inhibited the secretion of alpha-hemolysin at low concentrations. Real-time RT-PCR assay was further employed to evaluate the transcriptional level of hla, the gene encoding alpha-hemolysin, and RNAIII, an effector molecule of the agr system. The results indicated that the RNAIII expression and subsequent hla transcription were also inhibited by puerarin in a dose-dependent manner. Furthermore, puerarin significantly prevented human alveolar epithelial A549 cells from S. aureus-induced injury. Thereby, puerarin may be considered as a potential candidate for the development of antivirulence drugs in the treatment of S. aureus-mediated infections.

Establishment of Micellar Electro Kinetic Chromatography Detection Method for Puerarin in Pueraria Lobata(Willd.)Ohwi

Relative to the pharmacopoeia of high performance liquid chromatography (HPLC), establishing detection of puerarin in radix puerariae micellar electro kinetic chromatography (MEKC). Studying on buffer pH, borax concentration, separation voltage, SDS concentration of chromatographic conditions on the puerarin MEKC separation effect of influence, determined the best capillary electrophoresis conditions: test 250 nm wavelength, buffer of pH 9.0 0.1M KH2PO4solution-20 mM borax solution, separation voltage 18 kV, 40 mmol/L SDS concentration, 4.5 min retention time., grained puerarin MEKC standard equation: Y=36.522X-471.9, R2=0.9994(N=5), the puerarin concentration in 550 μg/mL to 750 μg/mL range comes out a good relationship. Get the lowest detection limit with 13.74 μg/mL. Puerarin content of the total isoflavone detected by MEKC was 45.35% that similar to 42.25% detected by HPLC. The method was stable, accurate, sensitive, possessed organic solvent consumption. The research results have applications in puerarin and its product quality evaluation system and promote isoflavonoids detection.

HP-β-CD-PLGA nanoparticles improve the penetration and bioavailability of puerarin and enhance the therapeutic effects on brain ischemia–reperfusion injury in rats

It is well known that puerarin attenuates ischemia-reperfusion injury and promotes function recovery of ischemic region. However, due to its reverse physiochemical properties, puerarin does not easily cross the blood-brain barrier. The aim of the present study is to create puerarin nanoparticles which increase and prolong the puerarin concentration in the brain. Using emulsion solvent evaporation techniques, we designed puerarin-loaded poly(D,L-lactic-co-glycolic acid) nanoparticles. Hydroxypropyl beta cyclodextrin (HP-β-CD) was used to increase the solubility of puerarin and gelatin to enhance viscosity of inner water phase, which improved puerarin entrapment. The drug release kinetics and nanoparticle degradation in phosphate buffered saline (PBS) were analyzed by electronic microscopy and high-performance liquid chromatography. Computerized tomography scans were used to detect the infarction volume and electroencephalogram (EEG) was recorded to estimate the recovery of brain function. The results showed that the combined HP-β-CD and gelatin significantly improved the entrapment efficiency. The infarction volume was significantly decreased on days 3 and 7 after the administration of puerarin nanoparticles compared with that of control and pure puerarin. EEG was also significantly improved. Puerarin nanoparticles are potentially applicable for the brain injury induced by ischemic-reperfusion.

Effect of pretreatment with puerarin on activation of LPS-induced RAW264.7 cells

To observe the effect of pretreatment with puerarin on activation of LPS -induced RAW264. 7 cells and secretory cytokines, and discuss its anti-inflammatory mechanism. Well-grown RAW264. 7 cells in the exponential phase were collected and randomly divided them into the blank control group, the LPS group and the puerarin pretreatment + LPS group. The cellular toxic effect of puerarin on RAW264. 7 cells was examined by CCK-8 assay, cell morphology was detected by Giemsa stain method, the changes in TNF-alpha and MIP-2 were tested by ELISA, and the expression of NF-kappaB p65 mRNA were determined by qRT-PCR. When puerarin was cultured with 1 mg x L(-1) LPS at a concentration of lower than 400 micromol x L(-1), it had not showed the cellular toxic effect (P < 0.05). Compared with the control group, the LPS group could significantly change the morphology of RAW264. 7 cells (increase in cell body, irregular shape, with a large number of pseudopodia extending). After intervention, the puerarin 100 micromol x L(-1) group could significantly inhibit LPS-induced cell morphological changes, while the puerarin 200 micromol x L(-1) and 400 micromol x L(-1) puerarin groups showed more notable inhibitory effects. However, there was no obvious difference between the two groups. The pretreatment with puerarin could inhibit the expression of TNF-alpha and MIP-2 in cell supernatant and NF-kappaB p65 mRNA in cells (P < 0.05). With increase in the puerarin concentration, its inhibitory effect gradually grew (P < 0.05), but did not reach the level of the blank control group. As a safe and effective natural anti-inflammatory drug, puerarin can significantly reduce the expression of inflammatory cytokines (TNF-alpha, MIP-2). Its mechanism may be related to the reduction of NF-kappaB p65 mRNA expression.

Osteoblast response to puerarin‐loaded porous titanium surfaces: An in vitro study

Recent studies demonstrate puerarin stimulates bone formation, suggesting its potential application in dental implantology field. The aim of this study was to investigate effects of puerarin-loaded titanium surfaces on the promotion of osteogenesis in preosteoblasts (MC3T3-E1). Puerarin was prepared onto titanium surfaces with varying concentration (10(-9) M, 10(-8) M, 10(-7) M, and 10(-6) M) by biomimetic calcium phosphate deposition process. Surface characteristics were performed by field-emission scanning electron microscope (FSEM), X-ray diffractometer (XRD), and Fourier transform infrared spectroscopy (FTIR). Puerarin concentration in the coatings was performed by High-performance liquid chromatograph (HPLC) analyses. FSEM observation showed puerarin concentration influenced formation of sharp flakes in the coating. The size of flakes decreased with increase of puerarin concentrations. XRD and FTIR examinations demonstrated the puerarin concentration did not affect the chemical compositions of coatings, which composed of octacalcium phosphate (OCP). Puerarin concentration on the surfaces of 10(-8) M group was 10.22 ± 0.32 ng/cm(2) . Puerarin had an increased effect on MC3T3-E1 ALP activities. Significant differences were found in 10(-8) and 10(-7) M groups on day 4, 10(-8) , 10(-7) , and 10(-6) M groups on day 7, and 10(-8) on day 14. In Type I collagen synthesis assay, 10(-9) and 10(-8) M on day 7, 10(-8) on day 14 showed significant differences compared with control group. Furthermore, this stimulatory effect of puerarin was also observed in osteocalcin release assay (p < 0.05, at 10(-8) M and 10(-7) M, maximal at 10(-8) M). These results indicate puerarin-loaded titanium surfaces promote accelerated osteogenic differentiation of preosteoblasts, which has the potential to improve the nature of osseointegration.

Flow Injection Chemiluminescence Determination of Puerarin in Pharmaceutical Preparations Using Eosin Y-Fenton System

A novel chemiluminescence (CL) system was established for the determination of puerarin in pharmaceutical preparations. It was shown that a strong CL signal was observed when Eosin Y reacted with the hydroxyl radicals which were generated from Fenton reagent in acidic medium. The CL intensity was decreased significantly when puerarin was added to the reaction system and partially scavenged the hydroxyl radicals in the solution. The extent of decrease in the CL intensity had a good stoichiometrical relationship with puerarin concentration. Based on this, a new method for the determination of puerarin using a flow injection chemiluminescence technique was developed. The experimental parameters that affected the CL intensity were optimized. Under the optimal conditions, the linear range for puerarin concentration was 8.0×10−8−2.0×10−6 mol/L (R=0.9982) with a detection limit of 7.5×10−9 mol/L (S/N=3) and the relative standard deviation was 1.7% for 4.0×10−7 mol/L puerarin (n=11). The proposed method was applied to the determination of puerarin in a puerarin injection with satisfactory results.

Electrocatalysis of Puerarin on a Nano‐CeO2/MWCNTs Composite Modified Electrode and Its Determination in Pharmaceutical Preparations

AbstractA good route for the fabrication of CeO2 nanoparticles (nano‐CeO2)/multi‐walled carbon nanotubes (MWCNTs) modified glassy carbon electrodes (GCE) was proposed. MWCNTs are used to immobilize nano‐CeO2. What′s more, with the addition of the MWCNTs, the agglomeration level of CeO2 nanoparticles can be reduced, the extremely large surface area can be obtained and the electron transfer rate can be increased. The morphological characterization of nano‐CeO2/MWCNTs was examined by scanning electron microscopy (SEM). The performances of the nano‐CeO2/MWCNTs/GCE were characterized with cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS) and typical amperometric response (i‐t). The potential utility of the constructed electrodes was demonstrated by applying them to the analytical determination of puerarin concentration. The catalytic oxidation of puerarin has a better result on nano‐CeO2/MWCNTs/GCE because of the synergistic effect of nano‐CeO2 and MWCNTs. An optimized limit of detection of 8.0×10−9 mol/L was obtained at a signal‐to‐noise ratio of 3 and with a fast response time (within 3 s). Additionally, the nano‐CeO2/MWCNTs/GCE exhibited a wide linear range from 0.04 to 6.0 μmol/L and high sensitivity.