Objective To develop a LC-MS/MS method for determination of daptomycinin human plasma.Methods The plasma samples were precipitated by acetonitrile,then the supernatant were separated on Eclipse Plus C18column (2.1 mm × 100 mm,3.5 μm) and eluted with acetonitrile-10 mmol · L-1 ammonium acetate (80∶ 20,containing 0.1％ formic acid),and the column temperature was 25 ℃.Detection of the analyte was achieved using positive ion electrospray ionization (ESI) in the multiple reaction monitoring (MRM) mode.The MS/MS ion transitions monitored were m/z 811.0→m/z 159.1 and m/z 237.1→m/z 194.1 for daptomycin and internal standard,respectively.The specificity,standard curve and lower limitation of quantitation,precision and recovery rate and stability as well as the matrix effect were investigated.Results The linear regression equation was y =1.49 × 10-2x-2.06 × 10-3 (r =0.9980) The linear range of daptomycin was 0.10-100 μg · mL-1,with the lower limit of quantitation (LLOQ) of 0.10 μg · mL-1.Accuracy was 97.1％-104.8％,Intra-day and inter -dayrelative standard deviations were both below 15％.The absolute recovery in plasma was81.9％-91.4％.The matrix effects were 90.01％-109.28％.Conclusion The established method israpid,sensitive,accurate,specific and reliable,and suitable for the determination ofdaptomycin in human plasma and pharmacokinetic study.

As an immediate-early protein of herpes simplex virus, infected-cell polypeptide 0 (ICP0) exhibits complicated interactions with host cells, and its regulatory function on gene expression is of great importance. Since the ICP0 encoding sequence contains many rare codons which are absent in E.coli, and ICP0 is highly unstable in prokaryotic cells, expression of entire ICP0 in prokaryotic cells has never been reported. In order to further investigate the function of ICP0, a recombinant plasmid was constructed by subcloning a cDNA fragment encoding an amino-terminal of 105 residues of the ICP0 protein into pGEX-5x-1 vector. The resulting GST-105 fusion antigen peptide was expressed with high efficiency in E.coli. Antibodies prepared after the immunization of mice with purified fusion protein can recognize not only the denatured ICP0 protein, but also the native ICP0 protein with normal biological conformation.

Objective To design a kind of bionic drag reduction needle and investigate its mechanism of painless injection. Methods Based on the analysis of mosquito's mouthpart structure, two types of bionic drag reduction injector needles, sawtooth needle and stripe needle, were designed with the painless feature as mosquito's blood-sucking action. The injection processes of normal smooth needle and two bionic drag reduction injector needles were simulated by using explicit kinetic software LS-DYNA, and analyzed the pain-relief mechanism of bionic drag reduction needles. Results Through contrast and analysis on the stress contour, internal energy curves and resistance curves of both the smooth needle and bionic needles during the puncturing process in soft tissue, the bionic needle was found to have obvious drag reduction effects. The drag reduction rate of the saw-tooth needle and stripe needle was 23.48% and 31.57%, respectively. Conclusions The bionic needles can minimize the friction resistance to achieve the painless injection by drag reduction, and its effect is determined by the structure and shape of the bionic needle.

Objective To evaluate the diagnosis values of whole blood IFN-γ release assay on pulmonary tuberculosis ( TB)and extra-pulmonary tuberculosis. Methods One hundred and eighty five patients with tuberculosis( including 119 pulmonary TB and 66 extra-pulmonary TB),139 patients with other respiratory diseases and 100 healthy people were enrolled. Mycobacterium tuberculosis infection testing was conducted with methods of IFN -γ release assay,sputum bacterial culture and sputum smear,respectively. Results of pathogen culture and/or clinical diagnosis were used as the golden standard to evaluate the sensitivity and specificity of these three methods. Results Compared with the results of clinical diagnosis,the sensitivity and specificity of IFN-γ release assays was 93. 51% and 84. 52%,respectively. There was no significant difference in sensitivity between pulmonary diagnosis( 90. 76%) and extra - pulmonary diagnosis (98. 49%)(P>0. 05). The sensitivity of sputum smear was 11. 76% in patients with pulmonary and 3. 03% in patients with extra - pulmonary. And the sensitivity of sputum bacterial culture in these two patient groups was 24. 37% and 3. 03%,respectively. Sensitivity of the IFN-γ release assay was higher than that of sputum culture and sputum smear (P<0. 05). The results of pathogen culture showed that 33 of 424 samples were positive,in which 2 were mycobacterium abscessus positive and 31 were mycobacterium tuberculosis positive. Compared with the results of pathogen culture,the sensitivity of IFN-γrelease assay was 90. 32%(95%CI:75. 10% -96. 65%). Conclusion IFN-γrelease assay is a fast,sensitive and convenient method to detect pulmonary and extra pulmonary tuberculosis. It is worthy to be applied to clinical practice.

Cell Adhesion ; Cell Movement ; Cell Proliferation ; Hep G2 Cells ; Humans ; Indoleamine-Pyrrole 2,3,-Dioxygenase ; genetics ; Transfection

AIMTo investigate the cardio-protective effect of crocetin on primary culture of cardiac myocyte treated with noradrenaline.

METHODSAfter adding crocetin, the primary culture of cardiac myocyte was injured by 1.0 micromol x L(-1) noradrenaline. The activity of lactic dehydrogenase (LDH), mitochondrion succinic dehydrogenase (MSDH) and ATPase were assayed. The mitochondrion membrane potential was detected by Rh123. The percentage of cardiac myocyte apoptosis was observed by flow cytometry.

RESULTSCrocetin significantly decreased the activity of LDH in culture supernatant, increased the activity of MSDH, ATPase (Na+-K+ ATPase, Ca2+ ATPase) and mitochondrion membrane potential.

CONCLUSIONCrocetin could alleviate the disturbance of energy metabolism and decrease the percentage of apoptosis of cardiac myocyte treated with noradrenaline.

Animals ; Animals, Newborn ; Apoptosis ; drug effects ; Calcium-Transporting ATPases ; metabolism ; Carotenoids ; isolation & purification ; pharmacology ; Cells, Cultured ; Crocus ; chemistry ; L-Lactate Dehydrogenase ; metabolism ; Membrane Potentials ; drug effects ; Mitochondria ; physiology ; Myocytes, Cardiac ; cytology ; enzymology ; Norepinephrine ; antagonists & inhibitors ; Plants, Medicinal ; chemistry ; Protective Agents ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Sodium-Potassium-Exchanging ATPase ; metabolism ; Succinate Dehydrogenase ; metabolism

OBJECTIVETo observe the effects of siwu tang on protein expression of bone marrow of blood deficiency mice and provide the theoretical and experimental basis for understanding the molecular mechanism of blood enriching function of siwu tang.

METHODBlood deficiency mice were established by using 3.5 Gy 60Co gamma-ray. With proteomic technologies including two-dimensional electrophoresis, image analysis, in-gel digestion, peptide mass fingerprinting and bioinformatics the proteins of bone marrow of blood deficiency mice were isolated, analyzed, and identified.

RESULTSiwu tang could reverse 10 up-regulated and 4 down-regulated proteins of blood deficiency mice bone marrow. Seven of the proteins were likely to be lymphocyte specific protein 1, proteasome 26S ATPase subunit 4, hematopoietic cell protein-tyrosine phosphatase, glyceraldehyde-3-phosphate dehydrogenase, growth factor receptor binding protein 14 and lgals12 respectively.

CONCLUSIONSiwu tang can regulate the protein expression of bone marrow of blood deficiency mice and thus promote the growth and differentiation of hematopoietic cells and then exert its effects on blood enriching function.

Adaptor Proteins, Signal Transducing ; Animals ; Drugs, Chinese Herbal ; pharmacology ; Female ; Medicine, Chinese Traditional ; Mice ; Mice, Inbred C57BL ; Phosphoproteins ; blood ; Proteins ; metabolism ; Proteome ; metabolism ; Proteomics ; methods ; Radiation Injuries, Experimental ; blood ; Whole-Body Irradiation

OBJECTIVETo investigate the biological difference of clonal cells between myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML).

METHODBone marrow (BM) clonal cells (which had cytogenetic markers detected by FISH assay) and blasts were quantitatively analysed in 51 MDS and 11 AML patients. The clonal cell percentage in orthochromatic normoblasts, granulocytes and megakaryocytes were assayed. The biological functions for phagocytosis and oxidation of MDS peripheral blood (PB) neutrophils were compared with that of normal controls.

RESULTSAlmost all MDS patients BM had a higher clonal cell percentage (mean 48.2%) than blasts percentage (mean 6.7%) (P < 0.01), but with the subtype of MDS advancing this percentage gap was closing up, and in 11 AML patients no such gap was observed. This gap in MDS patients with + 8 abnormality was smaller than in those with 5q -. In MDS BM, clonal cells were detected in segmented granulocytes (mean 45.9%), orthochromatic normoblasts (mean 46.0%) and mature megakaryocytes (mean 38.0%). In Addition, an approximate amount of clonal cells with the same karyotype abnormality in BM were detected in MDS PB (mean 37.3% in blood vs 48.6% in marrow). Functional analysis showed that the neutrophils in MDS PB could exert nearly normal physiological functions (P > 0.05), but those from AML could not as compared to healthy donors (P < 0.01).

CONCLUSIONThere is a significant difference in the biological features between MDS and AML clonal cells.

Adult ; Aged ; Aged, 80 and over ; Bone Marrow Cells ; pathology ; Cell Differentiation ; Clone Cells ; Female ; Humans ; Karyotyping ; Leukemia, Myeloid, Acute ; pathology ; Male ; Middle Aged ; Myelodysplastic Syndromes ; pathology

BACKGROUND: High-volume hemofiltration (HVHF) is technically possible in severe acute pancreatitis (SAP) patients complicated with multiple organ dysfunction syndrome (MODS). Continuous HVHF is expected to become a beneficial adjunct therapy for SAP complicated with MODS. In this study, we aimed to explore the effects of fluid resuscitation and HVHF on alveolar-arterial oxygen exchange, the Acute Physiology and Chronic Health Evaluation II (APACHE II) score in patients with refractory septic shock. METHODS: A total of 89 refractory septic shock patients, who were admitted to ICU, the Provincial Hospital affiliated to Shandong University from August 2006 to December 2009, were enrolled in this retrospective study. The patients were randomly divided into two groups: fluid resuscitation (group A, n=41), and fluid resuscitation plus high-volume hemofiltration (group B, n=48). The levels of O2 content of central venous blood (CcvO2), arterial oxygen content (CaO2), alveolar-arterial oxygen pressure difference P(A-a)DO2, ratio of arterial oxygen pressure/alveolar oxygen pressure (PaO2/PAO2), respiratory index (RI) and oxygenation index (OI) were determined. The oxygen exchange levels of the two groups were examined based on the arterial blood gas analysis at different times (0, 24, 72 hours and 7 days of treatment) in the two groups. The APACHE II score was calculated before and after 7-day treatment in the two groups. RESULTS: The levels of CcvO2, CaO2 on day 7 in group A were significantly lower than those in group B (CcvO2: 0.60±0.24 vs. 0.72±0.28, P<0.05; CaO2: 0.84±0.43 vs. 0.94±0.46, P<0.05). The level of oxygen extraction rate (O2ER) in group A on the 7th day was significantly higher than that in group B ( 28.7±2.4 vs. 21.7±3.4, P<0.01). The levels of P(A-a)DO2 and RI in group B on the 7th day were significantly lower than those in group A. The levels of PaO2/PAO2 and OI in group B on 7th day were significantly higher than those in group A (P<0.05 or P<0.01). The APACHE II score in the two groups reduced gradually after 7-day treatment, and the APACHE II score on the 7th day in group B was significantly lower than that in group A (8.2±3.8 vs. 17.2±6.8, P<0.01). CONCLUSION: HVHF combined with fluid resuscitation can improve alveolar- arterial-oxygen exchange, decrease the APACHE II score in patients with refractory septic shock, and thus it increases the survival rate of patients.

OBJECTIVETo study the technical feasibility of QAMS to determine different structural types of ingredients in Zhizi Jinhua pill, a Chinese patent medicine.

METHODEmodin was chosen as the internal reference substance. The relative correction factors (RCF) of berberine, baicalin, wogonoside, baicalein, aloe-emodin, wogonin, rhein, chrysophanol and physcion to emodin were calculated with the peak areas determined at 254 nm. The ruggedness of these RCFs was validated. The contents of emodin in the samples of Zhizi Jinhua pill were determined by using the external standard method, and the contents of the other nine ingredients were calculated by their RCFs. The contents of these ten ingredients in all samples were also determined by the external standard method.

RESULTNo significant differences were observed between the quantitative results of the two methods. The obtained RCFs were credible.

CONCLUSIONThe established QAMS method can be used for quantitative determination of different structural types of multi-compounds in Zhizi Jinhua pill. It can provide reference for the quality control of Chinese patent medicine.

Chromatography, High Pressure Liquid ; methods ; Dosage Forms ; Drugs, Chinese Herbal ; chemistry ; Quality Control