Quantitative analysis of multicomponents by a single marker and quality evaluation of Venenum Bufonis from different geographical origins.

Bufadienolides are the main bioactive components of Venenum Bufonis (VB) and have been widely used to treat different types of human cancers for decades. The bufadienolide content in VB varies significantly in materials from different geographical origins. In this work, a new strategy for the quality assessment of VB was developed through quantitative analysis of multi-components by single marker (QAMS). Cinobufagin was selected as the internal reference substance; seven bufadienolides were separated and simultaneously determined based on relative correction factors. The correlation coefficient value (r ≥ 0.9936) between QAMS and the normal external standard method proved the consistency of the two methods. According to the outcomes of 30 batches of VB samples, the contents of the seven bufadienolides were used for further chemometric analysis. All of the samples of VB from various geographical origins were divided into three categories based on hierarchical cluster analysis and radar plot, which indicated the crucial influence of geographical origins on VB. This study showed that QAMS combined with chemometristry could be used to comprehensively evaluate and effectively control the quality of VB from different geographical origins.

[Correlative study of bufogenin constituents in venom of Bufo bufo gargarizans].

As known,simultaneous determination of various chemical indicators is one of the future trends in quality control of traditional Chinese medicines because of the extremely complex chemical compositions. This project is to screen the quality markers that can accurately control the quality of the Bufonis Venenum by exploring the intrinsic correlation of components. In this study,venom of Bufo bufo gargarizans from 17 different sources were used as research samples,and the contents of 7 bufogenin were determined by HPLC-DAD. Then,the data obtained were analyzed by Spearman correlation analysis and principal component analysis( PCA). In addition,a stepwise regression analysis was used to establish a predictive model for the contents of the seven bufogenin components( independent variable) and the total contents of the bufogenin( dependent variable). The results indicated that there is a significant positive correlation between the contents of telocinobufagin and cinobufotalin,and there is a significant positive correlation between the contents of bufalin,cinobufagin and resibufogenin. In contrast,the contents of telocinobufagin and cinobufotalin are negatively correlated with the contents of bufalin,cinobufagin and resibufogenin. However,the correlation between gamabufotalin and bufotalin and other components are not obvious. Furthermore,further study found that there is a correlation between the sum of the contents of bufalin,cinobufagin and telocinobufagin and the total contents of the bufogenin. In fact,the application of bufalin,cinobufagin and telocinobufagin as the quality control indicators of the Bufonis Venenum can better reflect the quality characteristics of the Bufonis Venenum compared with the previous quality control indicators. The conclusions will provide a reference for the revision of the quality standards of the Bufonis Venenum.

Quantification of pantoprazole in human plasma using LC-MS/MS for pharmacokinetics and bioequivalence study.

A highly sensitive and rapid method for the analysis of pantoprazole in human plasma using liquid chromatography coupled to tandem electrospray ionization mass spectrometry was developed. The procedure involves a simple protein precipitation method with methyl alcohol and separation by RP-HPLC. Detection was performed by positive ion electrospray ionization in multiple reaction monitoring mode, monitoring the transitions m/z 384.1→200.0 and m/z 346.1→198.0, for quantification of pantoprazole and IS, respectively. The standard calibration curves showed good linearity within the range of 5-5,000 ng mL(-1). The lower limit of quantitation (LLOQ) was about 5 ng mL(-1). The extractive recovery of pantoprazole from the biological matrix was more than 77.58% and the matrix effect was complied with relevant provision. The intra-day accuracy of the drug containing serum samples was more than 92.19% with a precision of 0.79-5.36%. The inter-day accuracy was 85.49% or more, with a precision of 0.91-12.67%. Intra and inter-day accuracy of the assay at four concentrations were 97.9-98.2% with a precision of 4.2-13.9%. This method offered good precision and accuracy and was successfully applied to the pharmacokinetic and bioequivalence studies of 40 mg of enteric-coated pantoprazole in 20 healthy Chinese volunteers.

MicroRNA-6809-5p mediates luteolin-induced anticancer effects against hepatoma by targeting flotillin 1.

BACKGROUND: Luteolin (3,4,5,7-tetrahydroxy flavone) is a natural flavonoid abundant in fruits and vegetables. Although luteolin has shown pro-apoptotic activity in hepatocellular carcinoma (HCC) cells, the underlying molecular mechanism has not yet been clarified. PURPOSE: The aim of this study is to identify novel miRNAs involved in the action of luteolin in HCC cells and to explore the biological roles of these miRNAs. METHODS: The effect of luteolin on HCC cell growth was assessed using CCK-8 colony formation assay, flow cytometric analysis in vitro, and a xenograft model in vivo. miRNA expression profiles were assessed using next-generation sequencing. Differentially expressed miRNAs were validated by quantitative PCR. Bioinformatics analysis and luciferase reporter assay were utilized to confirm the binding of miR-6809-5p to the 3'-untranslated region (3'-UTR) of flotillin 1 (FLOT1). Furthermore, the effects of ectopic FLOT1 and miR-6809-5 expression on cell proliferation, colony formation, and cell apoptosis were also assessed. Western blotting analysis was used to detect activation of multiple signaling molecules including Erk1/2, p38, JNK, and NF-κB/p65 in the MAPK pathway. RESULTS: It was found that luteolin significantly inhibited HCC growth and caused apoptosis and cell cycle arrest at the G0/G1 phase in Huh7 cells, at the G2/M phase in HepG2 cells in vitro. Tumorigenic studies revealed that luteolin treatment significantly suppressed HCC growth in vivo. miR-6809-5p was upregulated by luteolin. Overexpression of miR-6809-5p suppressed HCC cell growth, while knockdown of miR-6809-5p reversed the anticancer effect of luteolin. With regards to its signaling mechanism, miR-6809-5p directly targets FLOT1in HCC cells. Enforced expression of FLOT1 prevented miR-6809-5p-mediated growth suppression. Downregulation of FLOT1 exerted growth-suppressive effects on HCC cells. Multiple signaling pathways including Erk1/2, p38, JNK, and NF-κB/p65 were inactivated by miR-6809-5p overexpression or FLOT1 downregulation. CONCLUSION: These findings indicated that miR-6809-5p mediates the growth-suppressive activity of luteolin in HCC, which is causally linked to FLOT1 downregulation. Induction of miR-6809-5p may provide therapeutic benefits in the treatment of HCC.

[Effects of acupuncture on behavior and striatal apoptosis in mice with Parkinson disease].

OBJECTIVE: To observe the effect of acupuncture of "Baihui" (GV 20), "Fengfu" (GV 16) and "Yanglingquan" (GB 34) on behavior and striatal apoptosis in Parkinson disease (PD) mice. METHODS: Twenty-eight adult male C57/ BL mice were randomly divided into normal control, PD model, acupuncture and medication (Levodopa and Benserazide Hydrochloride) groups. Acupuncture was applied to "Baihui"(GV 20), "Fengfu" (GV 16) and electroacupuncture (2 Hz, 1 mA) was applied to "Yanglingquan"(GB 34) for 15 min, once a day for 14 days. PD model was duplicated by intragastric administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The locomotor test, pole test and swimming test were conducted for assessing the animals' locomotion behavior before and after MPTP administration and after the treatment. The apoptotic neurons of the striatum were detected by terminal deoxyncleotidyl transferase-mediated dUTP-biotin nick end labeling assay. RESULTS: In comparison with the normal control group, the pole test score and apoptotic neuronal percentage were increased significantly (P<0.01), and the locomotor count, swimming test score were decreased remarkably in the model group (P<0.01). Compared with the model group, the locomotor counts and swimming test scores were increased obviously (P<0.05, P<0.01) and the pole test scores decreased apparently (P<0.05) in both acupuncture and medication groups after the treatment. No significant differences were found between the acupuncture and medication groups in the motor behavior (P>0.05). The effect of acupuncture was significantly superior to that of the medication in reducing neuronal apoptosis of the striatum (P<0.01). CONCLUSION: Acupuncture of "Baihui" (GV 20), "Fengfu" (GV 16) and "Yanglingquan"(GB 34) can improve PD mice's locomotor function, which is closely associated with its function in reducing neuronal apoptosis of the striatum.

[Characteristics of granular sludge of the internal circulation anaerobic expanded bed reactor].

Granular sludge characteristics of the Internal Circulation Anaerobic Expanded Bed (ICAEB) reactor is reported. Experimental results show that the mean COD removal was 89.0% and the internal circulation flow rate was about 12.5 when the reactor running steadily with the mean COD volumetric loading rate 28.2kg/(m3 x d). General granular sludge and suspended granular sludge are cultured in the reactor. The suspended granular sludge layer found in the fluid level of the second reaction section is dynamically stable. Keeping comparative steady operational condition could reduce the adverse influence of the suspended granular sludge layer, if necessary, suitable measurement could be adopted to remove the overabundant suspended granular sludge off.