OBJECTIVE: To purify and study Schwann cells cytoplasmic neurotrophic protein. METHODS: The dissociated SC taken from 300 newborn rats sciatic nerves were cultured, collected, ultrasonicated and ultraspeed centrifuged. The supernates were ultrafiltrated and concentrated by using ultrafiltration units with PM10, PM30, PM50 ultrafiltration membranes. The ultrafiltrated-concentrated solution with the protein molecular weight 10-30 ku, 30-50 ku and gt; 50 ku were collected respectively. The dissociated spinal cord motoneurons of 14 days embryonic rats were cultured with serum-free conditional medium and the additional SC cytoplasmic proteins were added into the medium. The results showed that the 10-30 ku and gt; 50 ku SC cytoplasmic proteins were able to maintain the survival of motoneurons for 24 hours. Then the 26 ku and 58 ku proteins were further extracted and purified from SC cytoplasm by high pressure liquid chromatography, and their neurobiological activities were studied. RESULTS: The 26 ku and 58 ku Schwann cell’s cytoplasmic proteins were able to maintain the survival of motoneurons cultured in the serum-free medium for 48 hours. The highest biological activity concentration is 20 ng per well. CONCLUSION: Schwann cells cytoplasm contains motoneuron neurotrophic proteins with molecular weight 26 ku and 58 ku.
ObjectiveTo study the effects of different penetration enhancers on the transdermal penetration of Miao medicine named Diploclisia affinis. MethodsImproved Franz diffusion cell was adopted as the apparatus for in vitro mouse' skin permeation. The kinetic parameters of percutaneous absorption, such as penetration rate, enhancing rate (ER), and lag time (Tlag) were determined by high performance liquid chromatography. Azone, oleic acid (OA) and borneol were investigated for percutaneous absorption effects. ResultsThe penetration rates of the medicine with 3% azone, OA and borneol added were respectively (214.1872±13.5690), (227.5544±9.8490), and (168.1187±21.5640) μg/(cm2·h), and the ER was 1.61, 1.71, and 1.26 compared with the penetration rate of that with nothing added. The Tlag was 2.1081, 1.8256, and 2.9655 hours. ConclusionAll the penetration enhancers can increase significantly the absorption of Miao medicine named Diploclisia affinis, especially 3% OA is the best, but 3% borneol may make the Tlag longer.
We in this study measured the site density of E-selectin in order to explore the practical pliability using radionuclide labeling method and γ-imaging of single photon emission computer tomography (SPECT). This method required labeling of antibody with 125I using Indogen method and binding of the labeled antibody to E-selectin. Labeled E-selectin was separated and purified in a Sephadex G25 column. The different fractions of the eluants were imaged, analyzed and quantified with SPECT method. For measuring the saturation curve of E-selectin, 130 μL of E-selectin solution with different concentrations were added in a 48-well plate and incubated overnight at 4℃. After incubation, 130 μL of labeled antibody solution were added and kept incubated for 30 min. The resulted mixture was washed, and the radioactivity in each sample was detected by SPECT. The levels of radioactivity were translated to site densities, and were used to plot a standard curve. The labeled product was quantitatively analyzed with SPECT. The labeling rate of E-selectin was 78%. The saturation curve of different concentration samples showed that when the concentration was in the concentration range of 0-1 mg/mL, the standard curve was y=6 045.7x—51.166, R2=0.997 9. Based on this finding, it could be concluded that γ-imaging is an important tool for analysis of radiolabeled product and determination of site density.
ObjectiveTo investigate the methodology of a newly developed highpressure liquid chromatography electrochemical system in urinary 8hydroxydeoxyguanosine (8OHdG) quantification. MethodsQuantification of urinary 8OHdG with ESA 5600 Coularray system among 21 children from liver cancer highrisk areas, Fusui country, in contrast with 63 controls from Nanning city, Guangxi province.ResultsThe resolution, sensitivity, linearity, precision and recovery of this approach all fully meet the requirement of routine urinary 8OHdG quantification. The mean 8OHdG level of this population was (5.26±0.41) ng/mg creatinine, higher than previous report 〔(4.62±0.091) ng/mg creatinine〕 by similar method.ConclusionWith cautious design and modification, this method is reliable and accurate in high throughput quantification of urinary 8OHdG.
Objective To establish a method for quality control of Astragalus Radix and Scutellariae Radix in Biqiaotong granules and provide basis for the establishment of quality standard. Methods The single-factor test method was used to investigate the factors of thin layer chromatography (TLC) conditions, including different extract method and solvents, developing system, comogemc agents, temperature, humidity, drawing amounts and thin layer boards, and to screen the best TLC conditions of Astragalus Radix and Scutellariae Radix . Results The TLC conditions of Astragalus Radix were used trichloromethane-methanel-water (13:7:2) as developing solvent, separated on silica gel G, heatd under 105℃ until the spots bacame clear. The TLC conditions of Scutellariae Radix were used methylbenzene-ethy acetate- formic acid-methanel (9:3:2:2) as developing solvent, separated on silica gel G, observed after 30 minutes under daylight until the spots were clear. Conclusions The spot features are clear, and with good separating degree, strong specificity, and good repeatability without the inference of negative control. The TLC method is simple, sensitive and accurate, which can be adopted for the quality control of Biqiaotong granules.
ObjectiveTo establish an accurate and sensitive high performance liquid chromatography-mass spectrometric (HPLC/MS/MS) method for determination of vorinostat (SHA) and M2 metabolites in human serum, which was applicable for pharmacokinetic study of SHA. MethodsThe essay was conducted with an API 3000 HPLC-MS/MS system consisted of a Gemini C18 column (50 mm×3 mm, 3 μm), and the mobile phase consisted of methanol-acetonitrile-water-1 mol/L NH3-formic acid (25:15:60:0.1:0.05) at a flow rate of 0.23 mL/min. Acidulated serum samples were extracted by 3.5 mL diethyl ether which contains 3% isopropyl alcohol and was operated under the multiple reaction monitoring mode using the electrospray ionization technique. d5-SHA was used as the internal standard. ResultsThe retention time of SHA, M2 metabolite and internal standard were 4.1, 3.1 and 4.0 minutes, respectively. The linear range of SHA and M2 metabolite were in the range of 1-1 000 and 2-2 000 ng/mL, and the limit of quantity were 1.0 and 2.0 ng/mL; the method recovery were 93.0%-99.3% and 88.11%-104.12%, respectively. Matrixes effect of SHA and M2 metabolite were blow 9.0%. ConclusionThis method for the quantitative determination of SHA and M2 in human serum was proved to be simple, rapid, sensitive and accurate; it can be applied in the determination of SHA and M2 metabolite.
ObjectiveTo understand the nutritional status of vitamin D in some children aged 0-14 in Mianyang during the past 3 years and the changes of vitamin D nutritional status under home protection during the coronavirus disease 2019 (COVID-19) epidemic, so as to provide a theoretical basis for the monitoring and reasonable supplementation of vitamin D in children in this area after the epidemic.MethodsThe clinical data of children aged 0-14 who underwent physical examination in the Children’s Health Department of Mianyang Central Hospital from January to April 2018, from January to April 2019 and from January to April 2020 were analyzed retrospectively. High performance liquid chromatography tandem mass spectrometry was used to detect vitamin D, including vitamin D2, vitamin D3 and 25-hydroxyvitamin D [25(OH)D] in children’s serum. The differences in vitamin D components and 25(OH)D between different genders, different age groups, and different years were analyzed.ResultsA total of 12 348 children were included. The average vitamin D2 was (4.89±6.02) ng/mL, the average vitamin D3 was (22.91±9.29) ng/mL, the average 25(OH)D was (27.81±10.53) ng/mL, and 9 434 cases had sufficient 25(OH)D. The differences in vitamin D2, vitamin D3, 25(OH)D and 25(OH)D nutritional status in 2018, vitamin D2 and 25(OH)D in 2019, and vitamin D2 in 2020 between different genders were not statistically significant (P>0.05). There were statistically significant differences in vitamin D3 and 25(OH)D nutritional status in 2019, vitamin D3, 25(OH)D and 25(OH)D nutritional status in 2020 between different genders (P<0.05). From 2018 to 2020, vitamin D2 was the highest in infant group (P<0.05), while vitamin D3, 25(OH)D and 25(OH)D nutritional status were the highest in children group (P<0.05); vitamin D2 (χ2=143.106, P<0.001) showed an overall downward trend, vitamin D3 (F=400.178, P<0.001) and 25(OH)D (F=447.384, P<0.001) showed an overall upward trend; 25(OH)D nutritional status (χ2=103.566, P<0.001) was the highest in 2019.ConclusionsThe overall vitamin D nutritional status of children in Mianyang area is acceptable. Under the home protection, the average level of children’s serum 25(OH)D has little change, while the nutritional status of 25(OH)D has decreased significantly. After the outbreak of COVID-19, more attention should be paid to the monitoring and supplementation of vitamin D in school-age female children.
ObjectiveTo explore the possible active mechanism of the basic fibroblast growth factor (bFGF) long circulation l iposome (LCL) (bFGF+LCL) on spinal cord traction injury in rats at the level of proteomics. MethodsTwenty Sprague Dawly rats were randomly divided into groups A and B, 10 rats in each group. The models of spinal cord traction injury was established at T12-L3 spines. The rats were not treated in group A, and the rats were treated with bFGF+LCL (20μg/ kg) in group B. At 3 weeks after operation, the rats were sacrificed for harvesting T13-L2 spinal tissue specimens. The protein was extracted and quantified in the spinal tissue firstly. The proteins from spinal tissue were separated by two-dimensional gel electrophoresis and identified by mass spectrometry. The different expression profiling was established in each group, and the differentially expressed protein was determined by comparing the level of each spot with gel imaging software and manually. The proteins were identified by nano ultra-high performance liquid chromatography-electrospray tandem mass spectrometry (NanoUPLC-ESI-MS/MS), and the proteins were classified. ResultsThe differentially expressed protein spots were found in 2 groups. Compared with group A, 4 spots were up-regulated and 6 were down-regulated in group B. NanoUPLC-ESI-MS/MS results showed that 18 significant proteins were identified in 26 differentially expressed proteins, including 4 apoptosis-related proteins, 3 nerve signal transduction related proteins, 7 proteins involved in metabolism, 1 unknown function protein, and 3 unnamed proteins. ConclusionThe differentially expressed proteins are found in spinal cord traction injury of rats treated with bFGF+LCL. bFGF+LCL can affect the proteins expression in rats with spinal cord traction injury. The possible active mechanism is that it has protective and repair effects on injured spinal cord by nerve signal transduction, and regulation of nerve cells apoptosis and metabolism.
ObjectiveTo determinate the concentration of isonicotinyl hydrazide (INH) in the hydrothorax and thereby increase the drug concentration of the hydrothorax to enhance the anti-tuberculosis efficacy through experiments. MethodsBetween February and June 2009, we used high performance liquid chromatography (HPLC) to determinate the concentration of INH in the hydrothorax of tuberculosis (TB) patients. The separation of sample was performed on Agilent ZORBAX SB-C18 (5 μm, 4.6 mm×2.5 mm) column with a fixed sample injection volume of 20 μL. The mobile phase was methanol-water (20︰80) at a flow rate of 1 mL/min. The ultraviolet detective wavelength was set at 254 nm; The column temperature was room temperature. ResultsIn the range of 0.25-10.00 μg/mL (r=0.998 9), moxi-floxacin showed a good linear relation in HPLC. The recoveries of moxi-floxacin at three concentrations (0.5, 5.0, and 10.0 μg/mL) were 97.95%, 100.64%, and 102.84%, respectively, with intra-day relative standard deviation (RSD) at 3.49%, 1.45%, and 2.03%, respectively and intra-day RSD at 3.85%, 5.68%, 4.15%, respectively. ConclusionThe measuring method adopted in the experiment can accurately determinate the concentration of INH in the hydrothorax with high sensitivity and exceptional reproducibility.
ObjectiveTo investigate the diagnostic value of lateral chromatography for cryptococcal capsular polysaccharide antigen in cryptococcal infection.MethodsThe data of patients who detected cryptococcal capsular polysaccharide antigen in West China Hospital of Sichuan University in 2018 were collected. The sensitivity, specificity, positive predictive value and negative predictive value of cryptococcal capsular polysaccharide antigen detected by lateral chromatography were analyzed. The samples with positive lateral chromatography and definite clinical diagnosis were compared with the results of ink staining and fungal culture.ResultsA total of 721 cases were detected. The sensitivity, specificity, positive predictive value and negative predictive value of lateral chromatography detection were 100.00%, 99.39%, 93.93%, 100.00%, respectively. The positive rates of ink staining, fungal culture and cryptococcal capsular polysaccharide antigen for cryptococcal infection were 63.46%, 48.00% and 100.00%, respectively. Sixty-two patients were clinically diagnosed, including 45 cases of cryptococcal meningitis (72.58%), 16 cases of cryptococcal pneumonia (25.81%), and 1 case of bone cryptococcal infection (1.61%).ConclusionsLateral chromatography detection for cryptococcal capsular polysaccharide antigen shows well performing sensitivity and specificity in the diagnosis of cryptococcal infection. Considering its rapid and simple pre-operation, cryptococcal capsular polysaccharide antigen detection with lateral chromatography has good application value for early diagosis of cryptococcal infection.