Objective To study the non-covalent binding between the tectoridin(TE) and plasma proteins.Methods The molecular weights of TE,human serum albumin(HSA),?1-acid glycoprotein(AAG) and the protein-drug complexes were measured by the electrospray ion trap mass spectrometry(ESI-MS).The maximum stoichiometric ratios were obtained according to the molecular weight change of the complexes before and after binding reaction.The binding constants(K) of the complexes were calculated by the Scatchard equation.The main sorts of binding force of the complexes were deduced according to the relationship between the reaction temperature and the thermodynamic parameters(?H and ?S).Results The K of the complexes were 1.914?104 mol/L for TE-HSA and 5.893?104 mol/L for TE-AAG,and the number of binding sites were 7.8 and 3.3,respectively.The main sorts of binding force between TE-HSA or TE-AAG were static-electricity gravitation.Conclusion ESI-MS is a good method for studying of the TE-protein non-covalent binding with some advantages in sensitivity,high-speed,and accuracy.

To study the influence of the compatibility of ophiopogonis tuber and Chinese magnoliavine fruit with gastrodia rhizome on the pharmacokinetics of gastrodin in rat, three dosages of compound Tianma granule extract (equivalent to gastrodin 50, 100, 200 mg x kg(-1)) and one dosage of Tianma extract (equivalent to gastrodin 100 mg x kg(-1) were administered to rats by intragastric administration separately. Plasma samples were collected at different times and treated with methanol and acetonitrile to precipitate protein. The contents of gastrodin in plasma were determined by HPLC method. The mean plasma concentration-time curves of different medication administration teams were processed with WinNonlin 5.2.1 pharmacokinetic software. The pharmacokinetic parameters of different medication administration teams were analyzed with SPSS statistics 17.0 software. The results indicated that the in vivo kinetic process of gastrodin was fitted to first-order absorption un-compartment model at low, middle dosages and zero-order absorption un-compartment model at high dosage of compound Tianma granule extract. By comparison with the pharmacokinetic parameters of gastrodin (100 mg x kg(-1)) in Tianma extract, the significant decrease for Cmax and significant increase for MRT0-infinity in compound Tianma granule extract indicated that the compatibility of ophiopogonis tuber and Chinese magnoliavine fruit with Gastrodia rhizome can delay the absorption, reduce the elimination rate and prolong the action time of gastrodin in vivo.

Triptolide (TP) is a major active component in Tripterygium root, but its therapeutic window was very narrow due to its severe multi-organ toxicity. In this work, the effect of TP combined with glycyrrhetic acid (GA) on mRNA expression and activity of four cytochrome P450 (CYP) enzymes in rat liver was studied after intragastric administration of TP (0.05, 0.3 and 0.6 mg x kg(-1) x day(-1)) and TP (0.6 mg x kg(-1) x day(-1)) combined with GA (30 mg x kg(-1) x day(-1)) for 7 consecutive days. Compared with the control, the high dose of TP significantly up-regulated the mRNA expression levels of CYP2E1, 1A2, 3A1 and 2C11, the co-administration of TP and GA further up-regulated the mRNA expression levels of CYP3A1, 2C11 and 2E1 as compared with the high dose of TP. Meanwhile, TP at high dose and combined with GA significantly increased CYP3A-associated testosterone 6beta-hydroxylation activity (2.2-fold and 4.1-fold, respectively) as compared with the control. Because TP is mainly metabolized by CYP3A2 in male rats, the present work indicated that TP-induced increase of CYP3A activity might be an important reason for the rapidly metabolic clearance of TP in rat liver, and GA can reduce the hepatotoxicity of TP by promoting its hepatic metabolic clearance. Furthermore, the results also suggest that the drug interactions might be occurred when TP and GA were co-administered with other CYP3A substrate drug.

Abstract: The activities of four CYP450 enzymes (CYP3A, 1A2, 2El and 2C) and the mRNA expression levels of CYP1A2, 2El, 2Cll and 3A1 in rat liver were determined after Wistar rats were orally administered with brucine (BR) at three dosage levels (3, 15 and 60 mg.kg-1 per day) and the high dose of BR combined with glycyrrhetinic acid (GA, 25 mg.kg-1 per day) or liquiritin (LQ, 20 mg.kg-1 per day) for 7 consecutive days. Compared with the control, brucine caused 24.5% and 34.6% decrease of CYP3A-associated testosterone 6beta-hydroxylation (6betaTesto-OH) and CYP2C-associated tolbutamide hydroxylation (Tol-OH), respectively, and 146.1% increase of CYP2El-associated para-nitrophenol hydroxylation (PNP-OH) at the high dose level. On the other hand, (BR+GA) caused 51.4% and 33.5% decrease, respectively, of CYP2El-associated PNP-OH and CYP1A2-associated ethoxyresorufin-O-de-ethylation (EROD) as compared with the high dose of BR group. Meanwhile, (BR+LQ) caused 41.1% decrease of CYP2El-associated PNP-OH and 37.7% increase of CYP2C-associated Tol-OH. The results indicated that the co-administration of BR with GA or LQ had effect on mRNA expression and activities of the CYP450 enzymes mentioned above to some extent, and the in vivo antagonism of LQ on BR-induced CYPs adverse effects and the in vivo inhibitory action of GA on CYP2E1 and 1A2 might play an important role in the detoxification of Radix Glycyrrhizae against Strychnos nux-vomica L.

Objective:To investigate the effects of Maijun'an Tablets on expression of endothelin-1(ET-1) and nitricoxide synthase(NOS) in tissues and plasma of spontaneous hypertensive rats (SHR). Methods:12 spontaneous hypertensive rats (SHR) were divided randomly into normal control group and drug administration group. Rats in drug administration group were orally given with Maijun'an Tablets (hydrochlorothiazide 10 mg?kg-1.d-1,puerarin 36 mg?kg-1.d-1 and total alkaloid of rhynchophylla 122 mg?kg-1.d-1) once a day for 4 weeks,and rats in control group were orally dosed with equal amount of physiological saline. All animals were sacrificed after 4 weeks. The mRNA and protein contents of ET-1 and NOS in plasma,liver,kidnay and blood vessel were determined. Results:Compared with the control group,Maijun'an Tablets can effectively reduce blood pressure of SHRs,regulate down the content of ET-1 and regulate up the content of NOS at mRNA and protein levels. Conclusion:The up-regulation of NOS and down-regulation of ET-1 at mRNA and protein levels were the major antihypertensive machnisam of Maijun'an Tablets for SHRs.

Background In recent years,some researches show that getting younger over the diabetes mellitus (DM) was an obrious trend.But studies of early diabetic retinopathy in young people with diabetes were few.Both patients and doctors did not put enough importance on it.Objective This study was to evaluate the earlier dynamic changes of retina morphology and function caused by type 2 DM in young population.Methods A cohort study was designed.One hundred and thirty-six young patients who were initially diagnosed as type 2 DM in Cangzhou Central Hospital were included from January to March in 2010 and 91 healthy individuals with the matched age were enrolled at the same period.Glycated hemoglobin (HbAlc) level in blood was assayed yearly,and ophthalmoscope,optical coherence tomography (OCT) and visual contrast sensitivity (CS) examinations were performed to evaluate the dynamic changes of retinal nerve fiber layer (RNFL) thickness around optical disc,retinal function and retinal microvessels of DM patients.In addition,the correlation between RNFL thickness or CS and average HbAlc level in DM patients was assessed.This study followed Declaration of Helsinki,and written informed consent was obtained from each subject.Results The levels of HbAlc in DM patients were (7.5 ±1.6) ％,(7.2 ±1.6) ％,(7.4 ±1.6) ％,(7.6±1.6)％ and (7.7 ±1.6)％ in 2010,2011,2012,2013 and 2014,respectively,and HbAlc≥7.0％ was observed in 74 patients (54.4％).The RNFL thicknesses of full optic disk and superior 1/4 quadrant of the optic disk were significantly declined in 2012,2013 and 2014 in the DM group compared with the normal control group (full disk:t =2.216,5.003,7.303;all at P ＜ 0.05.superior 1/4 quadrant:t =2.382,5.657,7.194;all at P ＜ 0.05),but no significant differences were found between them in 2010 and 2011 (all at P＞0.05).In the DM group,RNFL thicknesses of full optic disk were (99.94±10.01) μm and (96.32±10.32) μm in 2013 and 2014,which were significantly lower than (104.55±9.23) μm in 2010 (t=7.819,13.959,both at P＜0.05);the RNFL thicknesses of superior 1/4 quadrant around disk were (118.64± 13.98) μm and (115.06 ± 14.35) μm in 2013 and 2014,which were significantly lower than (127.48± 13.67) μm in 2010 (t =10.504,14.758,both at P＜0.05),However,the RNFL thicknesses were not significantly changed among the different time points in the normal control group (all at P＞0.05).In addition,the CS values of 3.0,6.0,12.0,18.0 c/d in 2013 and 2014 in the DM group were lower than those in the normal control group (all at P＜0.05),and the CS values in 2013 and 2014 were reduced in comparison with 2010 in the DM group (all at P＜0.05).Positive correlations were found between the differences of RNFL thickness or CS and the average HbAlc level in the DM group (r =0.640,F =8.991,P =0.000;r =0.541,F =7.572,P =0.000).Retinal microaneurysms were seen in two type 2 DM patients by ophthalmoscope at the end of follow-up.Conclusions The young people with type 2 DM have poor blood glucose control.The RNFLs are thinning and visual functions are declining in young type 2 DM patients with the lapse of disease course,and these retinal damages are associated with poor control of blood glucose.

Objective Adenosine receptor agonist NECA has a certain myocardial protection, but the specific mechanism is not clear.This paper aimed to study the effect and mechanism of NECA inhibiting endoplasmic reticulum stress to against myocardial ischemia reperfusion injury in rats.Methods 56 Wistar rats of SPF grade were selected and divided into Sham group, I/R group, NECA group and TUDCA group through random number table method.We established the isolated rat heart ischemia reperfusion model by using the Langendorff device.Sham group: heart threaded but not ligated, Kerb-Henseleit buffer continuous infusion 170min;I/R group: heart stability 20min, ischemia 30min, reperfusion 2h;NECA group and TUDCA group: heart stability 20min, ischemia 30min, reperfusion 2h, perfusion solutions containing 0.1μmol/L NECA and 30μmol/L TUDCA were respectively given at 5min before reperfusion, and ended at 30min after reperfusion.Transmission electron microscope was used to evaluate alterations of the myocardial ultrastructures.Western blot analysis was used to detected the expression levels of endoplasmic reticulum stress IREl-XBPl signaling pathway marker protein IRE1α, XBP1s.Immunohistochemical staining was used to detect the expression of IRE1α.Results The results of transmission electron microscopy showed that most of the myofilament ruptured, sarcomere contracture deformation, visible mitochondrial vacuoles degeneration in I/R group, and injury in NECA group and TUDCA group were less than the I/R group, appeared as the filaments arranged more neat, sarcoma only had mild contracture.Immunohistochemical results showed that IRE1αpositive staining was not found in the sham group, and the area of positive staining of IRE1α in I/R group was significantly increased, while the NECA group and TUDCA group were significantly decreased.Compared to the Sham group, the expression level of IRE1α and XBP1s was significantly increased in I/R group(P<0.05);but compared with the expression level of IREα and XBP1s in I/R group(1.72±0.27, 0.97±0.19), the NECA group(1.14±0.16, 0.6±0.13) and the TUDCA(1.07±0.27, 0.58±0.15) group were significantly decreased(P<0.05).Conclusion NECA can reduce endoplasmic reticulum stress through inhibiting IREl-XBPl pathway to protect the myocardium.

Objective:To scan the common differentially expressed genes in mice liver tissue following subacute exposure to hepatotoxicants,including alcohol,carbon tetrachloride,BCG & LPS,the alcohol extracts of Rhizoma Dioscoreae Bulbiferae and Senecio scandens Buch.-Ham.,and apply to the rapid toxic evaluation of TCM and other xenobiotics.Methods:The gene expression profiles of mice liver tissue,after respective administration of alcohol,CCl4,BCG associated with LPS,the alcohol extracts of Rhizoma Dioscoreae Bulbiferae and Senecio scandens Buch.-Ham.to healthy mice,were analyzed by use of mouse genome cDNA microarray.The correlations between the common differential expressed genes and the liver injury were analyzed based on the biological functions of those differentially expressed genes.Results:Among all of the five drug administration groups,there were 7 known function genes(BC05200,NM-019410,NM-173019,AB028272,AK088925,AK030862 and AK088816)and 1 unknown function gene(BC069871)differentially expressed.Beside AK088816,the other 7 genes were all down-regulate expressed.Conclusion:The common differentially expressed genes participate in the process of saccharometabolism,apoptosis,cell growth cycle,cytoskeleton and signal transmission,protein folding and protein ubiquitin.The abnormal expression of the common genes were closely correlated to the liver,and might be important for rapid hepatoxicity elucidation of traditional Chinese medicine and other xenobiotics.

Objective To develop the characteristic ESI-MS spectra of five Bulbus Fritillariae species and to be used for identification of Bulbus Fritillariae species.Methods The Bulbus Fritillariae medicinal materials were dipped in 18% ammonia solution,then the alkaloids were extracted by ultrasonic using the mixed solvent of ether-chloroform-ethanol(25∶8∶2.5).The extracts were analyzed directly by ESI-MS in positive ion mode.Results Bulbus alkaloids easily formed molecular ions(~+) in MS and dimerized easily in alcohol solution,which did not dimerize in acidity solution.Collision induced dissociation(S-CID) made these alkaloids easily lose a molecule of H_2O.The MS spectra of the extracts of five Bulbus Fritillariae species indicated that the major molecular ions were similar to each other,but different varieties showed very different relative abundances of these molecular ions.Conclusion The total alkaloid extracts of five Bulbus Fritillariae species have strong characteristic ESI-MS spectra and these spectra are of good reproducibility and precision,respectively.These characteristic spectra are useful for the identification of Bulbus Fritillariae species.