Six flavonol glycosides were isolated and calibrated from Ginkgo biloba extract, and then used to calibrate the content in 2 baiches of G. biloba reference extract, so was rutin. RSD values of rutin, kaempferol-3-O-rutinoside, kaempferol-3-O-rhamnoside-2-glu- coside, quercetin-3-O-rhamnop-yranosyl-2-O-(6-O-p-coumaroyl)-glucoside, kaempferol-3-O-rhamnopyranosyl-2-O-(6-O-p-coum-aroyl) - glucoside were around 1.1%-4.6%, nevertheless, RSD values of quercetin-3-O-glucoside and isorhamnetin-3-O-rutinoside were more than 5%. According to the results, the reference extract of G. biloba can be used as the substitute to determine rutin, kaempferol-3-O- rutinoside, kaempferol-3-O-rhamnoside-2-glucoside, quercetin-3-O-rhamnopyranosyl-2-O-(6-O-p-coumaroyl)-glucoside and kaempferol-3-0-rhamnopyranosyl-2-O-(6-O-p-coumaroyl)-glucoside instead of corresponding reference substances. So reference extract in place of single component reference in assay is feasible.
Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Flavonols ; chemistry ; isolation & purification ; Ginkgo biloba ; chemistry ; Glucosides ; chemistry ; isolation & purification ; Molecular Structure ; Spectrometry, Mass, Electrospray Ionization

AIM: To evaluate the implication of CXCL10-loop3-EGF fusion protein for the activities of targeting tumor and anti-angiopoiesis. METHODS: RT-PCR was preformed to amplify CXCL10 coding sequence from PBMC activated by IFN-γ. CXCL10-loop3-EGF fusion gene, which was conducted by Over-Lap Extention PCR, was hinged up with plasmid pTG19-T, transfected to E. coli DH5α and processed positive colony selection. After ligated with plasmid pET32a(+), recombinant CXCL10-loop3-EGF fusion gene was then transfected to E. coli Origami B (DE3) and induced to express its coding fusion protein his-CXCL10-loop3-EGF. The recombinant fusion protein CXCL10-loop3 -EGF was purified by His-bind affinity chromatograph, enterokinase cleavage, ultrafiltration and dislysis. The transwell chemotatic test and HUVEC angiopoiesis inhibition test were performed to determine the anti-tumor responses and anti-angiopoiesis activity of CXCL10-loop3-EGF fusion protein. RESULTS: CXCL10-loop3-EGF fusion protein was successfully constructed and confirmed by SDS-PAGE analysis and Western blotting. Significant PBMC chematatic activity and HUVEC anti-angiopoiesis activity were observed. CONCLUSION: CXCL10-loop3-EGF fusion protein, which has perfect anti-tumor activity, is successfully constructed.

Objective To construct the recombinant plasmid containing human filaggrin gene,purify and identify the immunoreactivity of the recombinant protein,and establish the indirect ELISA to detect AFA for diagnosis of RA.Methods The constructed plasmids were transformed into E. Coli Rosettagami(DE3).This fusion protein was purified by NAT chromatography.ELISA coated with the fusion protein Was established to detect the AFA in serum of patients,which included 114 cases of RA,56 cases of SLE,32 cases of OA and 40 cases of normal controls. The correlation between the results of AFA and anti-CCP in RA group were compared. Results 321 bp fragment of filaggrin gene was amplified and the recombinant expression vector pET-28a( + )-filaggrin was constructed. The sequence of filaggrin gene was the same as the sequence reported in the literatures. The Rosetta-gami (DE3) strains of E. Coli with recombinant vector showed high level of filaggrin protein after induction. The SDS-PAGE showed that the plasmid expressed the filaggrin fusion protein with molecule weight of 14 000 Da. The expression protein could be purified by Ni-NAT with activity. The absorbance value of AFA in RA group was 0.473 ±0. 248 while they were 0. 160 0. 088, 0. 121±0. 070, 0.050 0. ±018 in SLE, OA and normal groups respectively. There were significant differences of absorbance values of AFA between RA and SLE, OA, control group (t = 12.004, 14. 464, 18.078, P<0. 01, respectively). The positivities of anti-filaggrin in RA, SLE and OA were 48.2%, 5.4% and 3. 1% respectively. The positivities of AFA were significantly different between RA, OA and normal control groups (x~2 = 67. 088, P < 0. 01). There was positive correlation of results between AFA and anti-CCP antibody (r = 0.42, P < 0. 05 ) . The consistency rate of results between AFA and anti-CCP was 70. 1%. Anti-CCP was negative in 10 out of 114 patients with AFA positive. AFA can be used to diagnose RA with sensitivity of 48. 2% , specificity of 96.9% , positive predictive value of 93. 2% and negative predictive values of 67. 9% . Conclusions The purified human filaggrin fusion protein is successfully purified. The indirect ELISA method based on the recombinant protein shows good sensitivity and specificity. Joint detection with AFA and anti-CCP can improve the positive rate of detection.

s were 0.573±0.016, 0.707±0.008 and 0.711±0.013). Conclusions CXCL10 can express stablely in MCF-7 cell lines, which resulted in down-regulation of expression of VEGF and STAT3 gene. CXCL10 played an important role in anti-tumor effect.

Tetradrine-tashionone II(A)-PLGA composite microspheres were prepared by the SPG membrane emulsification method, and the characterization of tetradrine-tashionone II(A) -PLGA composite microspheres were studied in this experiment. The results of IR, DSC and XRD showed that teradrine and tashionone II(A) in composite microspheres were highly dispersed in the PLGA with amorphous form. The results of tetradrine-tashionone II(A) -PLGA composite microspheres in vitro release experiment showed that the cumulative release amounts of tetradrine and tashionone II(A) were 6.44% and 3.60% in 24 h, and the cumulative release amounts of tetradrine and tashionone II(A) were 89.02% and 21.24% in 17 d. The process of drug in vitro release accorded with the model of Riger-Peppas. Tetradrine-tashionone II(A) -PLGA composite microspheres had slow-release effect, and it could significantly reduce the burst release, prolong the therapeutic time, decrease the dosage of drugs and provide a new idea and method to prepare traditional Chinese medicine compound.
Benzofurans ; chemistry ; Benzylisoquinolines ; chemistry ; Drug Carriers ; chemistry ; Drug Compounding ; instrumentation ; methods ; Drugs, Chinese Herbal ; chemistry ; Kinetics ; Lactic Acid ; chemistry ; Microspheres ; Particle Size ; Polyglycolic Acid ; chemistry

Objective: To investigate the diagnose nutcracker syndrome with color Doppler flow imaging.Methods:Doppler sonographic findings in 47 children with orthostatic proteinuria or idiopathic hematuria and in 23 healthy control subjects were compared.The peak velocity,anteroposterior diameter and anteroposterior diameter ratio(hilar segment/aortomesenteric segment) were measured in the left renal vein.Results: The anteroposterior diameter ratio of the 2 segments were 4.13?2.34 in the patient group and 2.16?0.56 in the control.The anteroposterior diameter and the peak velocity in the aortomesenteric segment were(0.16?0.07)cm/s and(83.3?37.2)cm/s in the patient group and(0.24?0.09)cm/s and(43.1?10.8)cm/s in the control,with significant difference in between(P

Objective To investigate the mechanism of action of toll-like receptor 2 (TLR2),myeloid differentiation factor 88 (MyD88),and matrix metalloproteinase-9 (MMP-9) in cerebral ischemia-reperfusion injury in rats.Methods One hundred twenty-fiwe adult male SD rats were randomly divided into sham operation group,cerebral ischemia group and T2.5 (TLR2 antagonist) treatment group.A model of middle cerebral artery occlusion was induced by suture method.T2.5 (0.121 2 μg/g) was injected into jugular vein at the beginning of reperfusion in the T2.5 treatment group.Cerebral infarction volume,brain edema,bloodbrain barrier permeability and neurological deficit score were measured at 24 h after reperfusion.Western blotting was used to determine the expression levels of TLR2,MyD88 and MMP-9 at different time points in the ischemic cortex.Results Western blot analysis showed that compared with the sham operation group,the expression levels of TLR2 and MyD88 in the ischemic group increased significantly from 6 h after ischemia-reperfusion and lasted for 24 h (all P ＜ 0.05),while MMP-9 increased significantly at 24 h after ischemia-reperfusion (P ＜ 0.05).At 24 h after ischemia-reperfusion,the blood-brain barrier permeability,brain edema degree,cerebral infarction volume,and neurological deficit score in the ischemic group were significantly higher than those in the sham operation group (all P ＜0.05);at this time,the expression levels of TLR2,MyD88 and MMP-9 in the T2.5 treatment group were significantly lower than those in the ischemic group (all P＜ 0.05),and the blood-brain barrier permeability,brain edema degree,cerebral infarction volume,and neurological deficit score were significantly reduced (all P＜ 0.05).Conclusion TLR2,MyD88 and MMP-9 might be involved in cerebral ischemia-reperfusion injury.TLR2 antagonist T2.5 might inhibit the expression of MMP-9 through TLR2-MyD88 signaling pathway,thus alleviating cerebral ischemia-reperfusion injury.

Objective: To evaluate the safety of a herbal pilatory for external use. Methods : An acute eye irritation test were employed to detect the eye irritation of the herbal pilatory;a skin irritation test,a skin sensitization test and a skin phototoxicity test were employed to detect the dermal toxicity;Salmonella typhimurium reverse mutation assay（Ames test）and chromosome aberration test in CHL cells were employed to detect the effects of the pilatory on gene mutation and chromosome aberration in prokaryotic and eukaryotic cells. Results: When the eyes of rabbits exposed to the pilatory without rinse during the first 24 hours,the conjunctiva showed congestion and edema with the highest score of 2,corneal opacity was observed with the highest score of 1;however,these symptoms returned to normal within 72 hours,with the score reduced to 0. No irritation to the skin of rabits was found after exposed to the pilatory for fourteen days,no skin sensitization was introduced by Buehler test and no skin phototoxicity on guinea pigs was detected. There was no abnormal growth of reverse mutation colonies induced by the pilatory under S9 acitivation or not. There was no statistically significant rise of chromosome aberration rate in the exposed CHL cells compared to the control groups（P>0.05）. Conclusion Under the condition,the herbal pilatory showed mild and reversible irritation to eyes,while no dermal toxicity and genetic toxicity were observed. The safety of the herbal pilatory for external use is acceptable.

Objective: To observe the antiasthmaf ic, anti-inflammatory and immunological effects of Gubenchuansoukang Granule. Methods: The rats asthma model was established by ovalbumin sensitization. The model was identified by the study of the asthma incubation period and the total white cells counts in bronchoalveolar lavage fluid (BALF). The anti-inflammatory effect was observed by the acute inflammation and chronic inflammation induced xylene and agar. To observe the effect of Gubenchuansoukang Granule on humoral immunity and cellular immunity induced by chicken red blood cell and 2, 4-dinitrochlorobenzene. Results: Gubenchuansoukang Granule could lengthen the asthma incubation period, reduce the quantity of total white cells in BALF, against the acute inflammation and chronic inflammation, and restrain the humoral immunity and cellular immunity. Conclusion: Gubenchuansoukang Granule had antiasthmafic, anti-inflammatory and immunosuppression effects.

Objective:To investigate the changes of c-Jun N-terminal kinase (JNK) and matrix metalloproteinase-9 (MMP-9) expressions after cerebral ischemia-reperfusion in rats and whether JNK inhibitor SP600125 can protect the cerebral ischemia-reperfusion injury by down-regulating the expression of MMP-9. Methods:Eighty-five adult male SD rats were divided into sham-operated group ( n=35), ischemia group ( n=35) and SP600125 group ( n=15). The middle cerebral artery occlusion models in rats of ischemia group and SP600125 group were established by thread embolism method, and reperfusion was performed one h after ischemia. Rats in the SP600125 group were injected with SP600125 solvent (being dissolved in 10% dimethyl sulfoxide with final concentration of 20 mmol/L) in the lateral ventricle 30 min before cerebral ischemia. At 48 h after reperfusion, neurological impairment scale scores, volume ratio of ischemic lesion, and moisture content of brain tissues in rats from each group were evaluated. The protein expression levels of phosphorylated (p)-JNK, JNK and MMP-9 in the ischemic cortex were detected by Western blotting in rats from the sham-operated group and ischemic group at 3, 6, 24 and 48 h after reperfusion, and in rats from SP600125 group at 48 h after reperfusion. Results:As compared with the sham-operated group, rats in the ischemia group had significantly decreased neurological impairment scale scores, and significantly increased infarction volume ratio and moisture content of brain tissues 48 h after reperfusion. At 24 and 48 h after reperfusion, the p-JNK/JNK values and MMP-9 protein level in the ischemia group were significantly increased as compared with those in the sham-operated group ( P<0.05). As compared with the ischemia group, at 48 h after reperfusion, rats in the SP600125 group had significantly increased neurological impairment scale scores, and significantly reduced volume ratio of ischemic lesion and moisture content of brain tissues, and significantly reduced p-JNK/JNK values and MMP-9 protein level in the ischemic cortex ( P<0.05). Conclusion:The expressions of JNK and MMP-9 are increased after cerebral ischemia-reperfusion, and the JNK inhibitor SP600125 may decrease MMP-9 expression to reduce the degrees of cerebral infarction after cerebral ischemia-reperfusion, thereby playing a role in brain protection.