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.
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.
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.
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 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 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 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.
High performance liquid chromatography (HPLC) is currently the mainstream technology for detecting hemoglobin. Glycated hemoglobin (HbA1c) is a gold indicator for diagnosing diabetes, however, the accuracy of HbA1c test is affected by thalassemia factor hemoglobin F (HbF)/hemoglobin A2 (HbA2) and variant hemoglobin during HPLC analysis. In this study, a new anti-interference hemoglobin analysis system of HPLC is proposed. In this system, the high-pressure three-gradient elution method was improved, and the particle size and sieve plate aperture in the high-pressure chromatography column and the structure of the double-plunger reciprocating series high-pressure pump were optimized. The system could diagnose both HbA1c and thalassemia factor HbF/HbA2 and variant hemoglobin, and the performance of the system was anti-interference and stable. It is expected to achieve industrialization. In this study, the HbA1c and thalassemia factor HbF/HbA2 detection performance was compared between this system and the world’s first-line brand products such as Tosoh G8, Bio-Rad Ⅶ and D10 glycosylated hemoglobin analysis system. The results showed that the linear correlation between this system and the world-class system was good. The system is the first domestic hemoglobin analysis system by HPLC for screening of HbA1c and thalassemia factor HbF/HbA2 rapidly and accurately.
Objective To determine the best threshold value of hemoglobin A2 (HbA2) for diagnosis of β-thalassemia (β-thal) carriers by using high performance liquid chromatography (HPLC), and to improve the application value of HbA2 as a diagnostic index for β-thal carriers to reduce the rates of missed diagnosis and misdiagnosis. Methods Using reverse dot blot (RDB) as a gold standard method, HbA2 results of 1 007 β-thal carriers and 606 normal controls in the past two years determined by HPLC were divided into true positive, false positive, true negative, and false negative based on the different threshold values of HbA2 results. Then, the evaluation indexes such as sensitivity, specificity, positive and negative likelihood ratio, and Youden’s index were evaluated. Next, the receiver operator characteristic (ROC) curve was drawn to determine the best threshold value of HbA2 for diagnosis of β-thal carriers by HPLC. Results If ≥4.0% was taken as the threshold value of HbA2 for diagnosis of β-thal carriers by HPLC, the evaluation indexes values were shown as follows: sensitivity 99.21%, specificity 99.34%, positive likelihood ratio 150.30, negative likelihood ratio 0.008, and Youden’s index 0.99. The Youden’s index was better than the other threshold values, and the corresponding tangent point was the peak point of the ROC curve. Conclusion When ≥4.0% serves as the best threshold value of HbA2 for diagnosis of β-thal carriers using HPLC, integrated evaluation performance of the corresponding sensitivity and specificity is the most ideal, and the authenticity of the diagnostic test is the best.
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.