Objection To estabish a HPLC methold for determining the contents of pinoresinol diglucoside and chlorogenic acid in Sam Soup and its solid decoction. Methods Chromatographic column was Ultimate XB-C18 reversed-phase chromatography column (250 mm×4.6 mm, 5 μm). Mobile phase was acetonitrile-0.4% glacial acetic acid with gradient elutim. The flow rate was 1.0 mL/min, detection wavelength was 277 nm, and column temperature was 40 ℃. Results The pinoresinol diglucoside and showed good linear relationship in the range of 5.0-50.0 μg, r =0.999 9 (n =6), and chlorogenic acid showed linear relationship in the range of 5.4-54.0 μg, r=0.999 6 (n=6). The average recoveries of pinoresinol diglucoside and chlorogenic acid were 98.75% (RSD=1.39%) and 101.11% (RSD=2.69%), respectively. Conclusion The established method was accurate and reliable. The contents of pinoresinol diglucoside and chlorogenic acid in solid decoction are higher than traditional decoction of Sam Soup.

To examine the effects of ischemia and anoxia on cell activation and cell cycle of astrocytes in vitro, the cell cycles and the proliferation of astrocytes in different time points after ischemia and anoxia were studied by flow cytometry and BrdU labeling and the expression of GFAP and cyclin D1 was detected by the fluorescence immunochemistry. After ischemia and anoxia in vitro, the astrocytes in S phase were significantly increased as compared with those in the normal group and the proliferating ability of the astrocytes was highest 6 h after the treatment as revealed by BrdU pulse labeling, but the astrocytes in S phase and proliferating ability were decreased after 6 h. At the early stages of ischemia and anoxia, the positive staining intensity of GFAP was increased, peaked at 6th h, while 12 h after the ischemia and anoxia, the positive staining intensity of GFAP became weak, and the expression of cyclin D1 was gradually increased after the ischemic and anoxic damage. It is concluded that astrocytes are activated to proliferate and enter new cycle events by ischemia and anoxia, and cyclin D1 is implicated in the proliferation and repair of astrocytes. The cell cycle events are closely associated with the proliferation and activation of astrocytes.
Animals, Newborn ; Astrocytes/*cytology ; Cell Cycle ; Cell Hypoxia ; Cell Proliferation ; Cells, Cultured ; Cyclin D1/biosynthesis ; Cyclin D1/genetics ; Glial Fibrillary Acidic Protein/*biosynthesis ; Glial Fibrillary Acidic Protein/genetics ; Rats, Wistar

Objective To evaluate the flexibility and efficacy of the Willis covered stent in the treatment of distal internal carotid artery(DICA)aneurysms.Methods The study was approved by the anthors' institutional review board,and the research was conducted by the authors' institution and the MicroPort Medical Company(Shanghai,China).Thirty-one patients with 33 DICA aneurysms were considered for treatment with a Willis covered stent.The angiographic assessments were categorized as complete or incomplete occlusion.The data on technical Success,initial and final angiographic results,mortality,morbidity,and final clinical outcome were collected,and follow-up was performed at 1,3,6,and 12 months and yearly after the procedures.Results Navigation and deployment of the covered stents were succssfnl in 97.6%(41 of 42;95%confidence interval[CI]:93%,102%)of the attempted stent placement procedures.The initial angiographic results showed a complete occlusion in 23 patients with 25 aneurysms(of 32 aneurysms,78.1%[95%CI:63%,93%])and an incomplete occlusion in seven patients with seven aneurysms(21.9%).The angiographic follow-up(mean,14 months[95% CI:12,15 months])findings exhibited a complete occlusion in 27 patients with 29 aneurysms(of 31 aneurysms,93.5%[95%CI:84%,103%])and an incomplete occlusion in two aneurysms(6.5%),with a mild in-stent stenosis in two patients.The clinical follow-up(mean,27 months[95% CI:23,30 months])demonstrated that 15 patients experienced a full recovery and 14 patients improved.Conclusion The preliminary results demonstrate good flexibility and efficacy of the Willis covered stent in the treatment of DICA aneurysms in selected patients:longer follow-up and expanded clinical trials are needed.

Antithrombin III (AT III) is the most important anti-clotting substance. Recombinant human antithrombin III (rhAT III) expressed in transgenic goat milk attracts more and more attention. Develop an effective purification route for rhAT III is vital to its industrial production. An efficient purification method was developed for the rapid purification of rhAT III by isoelectric precipitation and heparin affinity chromatography. First, casein was effectively removed by isoelectric precipitation. rhAT III was further purified by heparin affinity chromatography. In the process of heparin affinity chromatography, the effects of pH and temperature on the stability of rhAT III were studied, and the effects of operating conditions, elution gradient, flow rate and sample loaded, on the purification efficiency were also studied. Under the optimized conditions, the protein recovery of rhAT III was about 90% with purity over 99%, while its activity recovery was about 50%. Such a purification process is very simple and effective, and it would provide a valuable reference for the further scaling-up of industrial production.
Animals ; Animals, Genetically Modified ; Antithrombin III ; biosynthesis ; Chromatography, Affinity ; Female ; Goats ; Heparin ; Humans ; Mammary Glands, Animal ; metabolism ; Milk ; chemistry ; Recombinant Proteins ; biosynthesis

Objective To measure the dose to the primary operator and assistant operators by employing eight beam projections commonly used in coronary angiography with and without radiation protection shields in order to supply helpful guidance on radiation protection in cardiac intervention.Methods From 20 to 180 cm above the ground at the primary and assistant operators' locations,a DoseAware personal dose meter was placed in terms of an increment of 20 cm to measure radiation dose.Eight commonly used beam projections were performed,including LAO (left anterior oblique) 45°,RAO (right anterior oblique) 30°,CRAN (cranial)25°,cranial LAO (LAO45°/25°),caudal LAO(LAO45°/ 25°),CAUD (caudal) 25°,cranial RAO (RAO30°/25°),caudal RAO (caudal RAO30°/25°).Under the two different conditions,with or without radiation protection shields,the doses to the operators in the selected beam projections were respectively recorded at nine measuring positions and the shielding factor were calculated.Results The primary operator was effectively protected with radiation protection shields.In the standing area of the primary operator,except for the position at the height of 120 cm (radiation dose rate:0.35-4.78 mSv/h;shielding factor:27.67％-89.33％),the shielding factor for each measuring position was above 91％.Higher radiation doses were found at caudal LAO,LAO,and cranial LAO.The shielding factor for the assisting operator was lower than for the primary operator.In the standing area of the assisting operator (radiation dose rate:0.27-1.86 mSv/h;shielding factor:30.34％-92.13％),the peak levels were found at the height of 80,100,140 cm.And caudal RAO,caudal LAO,CRAN,LAO were found to have received higher radiation doses.Conclusions Emphasis should be attached to the use of radiation shields in coronary angiography.With radiation protection shields,higher dose is still recorded in caudal LAO,LAO,cranial LAO,caudal RAO.Furthermore,it should be paid more attention to radiation protection at 80-140 cm height,and less prolonged exposure should be employed in those beam projections mentioned above.

Objective To evaluate the radiation protective efficacy of different types and the positions of ceiling-suspended lead shield to the principal and assistant interventional operators in order to provide a scientific basis for the selection of optimal scheme in using ceiling-suspended lead shield.Methods At the principal and assistant interventional operators’ standing places the personal dose-measuring instruments were set up, which were placed at the height of 20 cm to 180 cm above the ground with an interval distance of 20 cm between each other.The postero-anterior (PA) projection and left lateral projection were used.The ceiling-suspended protection lead shields included lead glass (glass type) and lead glass with connected lead flexible stripe below (mixed type).The placed sites of the protection lead-shields were close to the principal operator, away from the principal operator, on the left side of the principal operator and close to the X-ray tube respectively.The radiation doses of PA projection and left lateral projection were determined.The real-time radiation dose rate and dose shielding rate at the nine measuring positions for the principal operator and assistant operator were separately calculated.The results were analyzed.Results The radiation protection of the glass type was slightly superior to that of the mixed type, but the difference was not significant.The principal operator was best protected when the shield was positioned close to him in the PA projection, and for left lateral projection the principal operator was best protected when the shield was positioned on his left side.For the assistant operator, the optimal protection was obtained when the shield was positioned close to him in both PA and left lateral projection.In the optimal position of ceiling-suspended lead shield, the highest radiation dose rate (0.71 mSv/h in glass group and 1.07 mSv/h in mixed group) was recorded on the principal operator at the height of 120 cm at PA projection, and higher radiation dose rate (≥0.47 mSv/h) was recorded on every point of both operators at the left lateral projection.Meanwhile, the overall received radiation doses of the two groups were very close.At the principal operator standing area, except for the position of 120 cm height (attenuation ratio 60.11% in glass group and 39.89% in mixed group), the attenuation ratio of each measuring point was above 93%.And the assistant operator standing area the attenuation ratio was 57%-97%.The lateral shielding ratio was generally slightly higher than PA shielding ratio.Conclusion The radiation protection effect of the two type shields is quite similar, both shields can obtain excellent protection efficacy.But the radiation dose at the height of 120 cm above the ground at PA projection is higher for the principal operator, while at lateral projection the radiation dose at all height levels is still relatively higher for both operators.Therefore, the radiation protection at the level of 120 cm height needs to be strengthened and the lateral projection exposure should be used as less as possible.

As a virus-like particle, hepatitis B surface antigen (HBsAg) was the primary component of hepatitis B vaccine. HBsAg was maintained by the non-covalent interaction of proteins and lipids. The intact structure of HBsAg particle was vital to its function. However, there was no report about the effects of solvent environment on HBsAg structure. In this paper, we studied the effects of temperature, pH, ionic type and salt concentration on HBsAg structure. The results showed that HBsAg was stable at normal temperature, but began to denature above 60 degrees C. The aggregation of HBsAg at pH 3.0 and 4.0 was nearly irreversible, but partly reversible at pH 5.0. The influence of ionic type on HBsAg was generally in accordance with Hofmeister sequence, except that SO4(2-) caused more aggregation than F-. HBsAg aggregates started to be visible in 0.4 mol/L (NH4)2SO4, and the extent of aggregation increased with the salt concentration. Therefore, caution must be taken when using (NH4)2SO4 in the hydrophobic chromatography purification of HBsAg.
Ammonium Sulfate ; chemistry ; Hepatitis B Surface Antigens ; chemistry ; Hepatitis B Vaccines ; chemistry ; Hepatitis B virus ; chemistry ; Hydrogen-Ion Concentration ; Protein Denaturation ; Solvents ; Temperature

Endotoxin removal is essential for the safety of biological products. To remove endotoxin efficiently, we used polymyxin B (PMB) affinity adsorbent to remove endotoxin from protein solutions by static adsorption. We studied the effects of spacer length and ligand density of the affinity adsorbent, pH, salt type and concentration, protein type and concentration, endotoxin concentration, and additive on endotoxin removal and protein recovery. Endotoxin content and protein concentration were determined by test and Lowry assay respectively. The results showed that PMB affinity adsorbent had high capacity, high adsorption speed, high removal efficiency and good reusability. In addition, ligand density, pH, salt concentration and the isoelectric point and hydrophobicity of protein all had remarkable influence on the endotoxin removal. Under the optimal conditions, the recoveries of hemoglobin, human serum albumin and lysozyme were 87.2%, 73.4% and 97.3%, respectively, and the corresponding endotoxin removal rates 99.8%, 97.9% and 99.7%, respectively. This study illustrated the effects of solution conditions on the efficiency of endotoxin removal and protein recovery, and would provide useful reference for the efficient removal of endotoxin from biological products.
Adsorption ; Chromatography, Affinity ; methods ; Drug Contamination ; prevention & control ; Endotoxins ; isolation & purification ; Hemoglobins ; isolation & purification ; Polymyxin B ; chemistry ; Proteins ; isolation & purification ; Pyrogens ; isolation & purification ; Serum Albumin ; isolation & purification ; Solutions

Novel ion exchange adsorbents were synthesized by immobilizing sulfopropyl derivative onto homemade highly cross-linked agarose beads. The effects of different ligand densities (from 0.05 to 0.24 mol/L) on static and dynamic adsorption of the adsorbents were investigated using lysozyme as a model protein. Based on these results, rHLF was purified from the transgenic milk by our SP media. 1 mL high density (0.24 mol/L) adsorbent could handle 50 mL rHLF-containing milk. The mass recovery of rHLF was 86.5% and the purity was 98.5%. CD spectra demonstrated that the native structure of rHLF was not affected in the purification process. The biological functions of the purified rHLF, including iron binding, releasing and antimicrobial activities were then investigated. The results showed that rHLF had comparable iron binding and releasing activity to that of native HLF. 5 g/L concentration of rHLF significantly inhibited the growth of Escherichia coli. These studies lay a solid foundation for the wide application of our self-prepared ion exchange adsorbents in protein purification.
Animals ; Cattle ; Female ; Humans ; Lactoferrin ; biosynthesis ; genetics ; Mammary Glands, Animal ; metabolism ; Milk ; chemistry ; Milk, Human ; chemistry ; Recombinant Proteins ; biosynthesis ; genetics ; isolation & purification

To examine the effects of ischemia and anoxia on cell activation and cell cycle of astrocytes in vitro, the cell cycles and the proliferation of astrocytes in different time points after ischemia and anoxia were studied by flow cytometry and BrdU labeling and the expression of GFAP and cyclin D1 was detected by the fluorescence immunochemistry. After ischemia and anoxia in vitro,the astrocytes in S phase were significantly increased as compared with those in the normal group and the proliferating ability of the astrocytes was highest 6 h after the treatment as revealed by BrdU pulse labeling, but the astrocytes in S phase and proliferating ability were decreased after 6 h.At the early stages of ischemia and anoxia, the positive staining intensity of GFAP was increased,peaked at 6th h, while 12 h after the ischemia and anoxia, the positive staining intensity of GFAP became weak, and the expression of cyclin D1 was gradually increased after the ischemic and anoxic damage. It is concluded that astrocytes are activated to proliferate and enter new cycle events by ischemia and anoxia, and cyclin D1 is implicated in the proliferation and repair of astrocytes. The cell cycle events are closely associated with the proliferation and activation of astrocytes.