To develop an analytic method for qualitative and quantitative analysis of dodecatetraenamides A, B in 42 samples of two official species of Asari Radix et Rhizoma( ARR) (37 samples of Asarum heterotropoides var. mandshuricum with different collection time and 5 samples of Asarum sieboldiivar. seoulense). The HPLC-IT-TOF-MS/MS methods for the qualitative and UPLC-PDA methods for the quantitative analysis were established. Dodecatetraenamides A, B were identified by comparing the retention time, UV absorption spectrum and quasi-molecular ion peak [ M + H]+ with the reference compound using HPLC-IT-TOF-MS/MS. The content of dodecatetraenamides A and B in ARR were determined by UPLC-PDA. The separation was successfully carried out on a ACQUITY UPLC BEH C18 (2.1 mm x 100 mm, 1.7 µm) column eluted with mobile phases of water (A) and acetonitrile (B) in gradient program (0-3 min, 35% B; 3-5 min, 35%-36% B; 5-6 min, 36%-43% B; 6 min-11 min 43% B; 11-12 min, 43%-100% B). The column temperature was 45 °C, and the detection wavelength was set at 254 nm. The flow rate was 0.6 mL · min(-1). On one level mass spectrometry scanning, the results showed that the quasi-molecular ion [M + H] + of both dodecatetraenamides A and B were m/z 248.20. The quantitative method with UPLC-PDA has made the baseline separation of the constituents, which were reported as mixtures in the most literatures. The average recovery of dodecatetraenamides A and B were 97.90% and 99.86%, the relative standard deviation were 0.4% and 1.1%, respectively. The contents of dodecatetraenamides A, B in all ARR samples was in the range of 0.11-3.89 and 0.24-6.65 mg · g(-1). Their contents reduced with the extension of storage time. Compared with the samples of 2013, the average content of the two constituents in the samples collected in year 2002-2003 reduced 34% and 36%, respectively (P < 0.05). Compared the A. sieboldii var. seoulense and A. heterotropoides var. mandshuricum with the same collective time and production area, the average contents of the two constituents in latter were up to (1.59 ± 0.75) mg · g(-1) and (2.90 ± 1.17) mg · g(-1), respectively, significantly higher than that in A. sieboldii var. seoulense (dodecatetraenamide A were (0.78 ± 0.52) mg · g(-1), dodecatetraenamide B were (1.69 ± 0.83) mg · g(-1)) (P < 0.05). The content of the dodecatetraenamide A in overground part was in the range of 0.11-0.33 mg · g(-1), dodecatetraenamide B was 0. 24-0.60 mg · g(-1), which were much lower than that of the underground part of ARR (dodecatetraenamide A was in the range of 0.73-3.89 mg · g(-1), dodecatetraenamide B was 2.11-6.24 mg · g(-1)). The method was certified to be simple, accurate and reliable and could be used for qualitative and quantitative analysis of dodecatetraenamide A and B in different species of ARR, also can be used for the comprehensive quality control of traditional Chinese medicine, Asari Radix et Rhizoma.
Amides ; chemistry ; Asarum ; chemistry ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Mass Spectrometry ; Molecular Structure ; Rhizome ; chemistry

OBJECTIVETo analyze and identify the principal composition of ethyl acetate part of Huangqi Danggui decoction by high performance liquid chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS).

METHODThe analysis conditions are as follows: Zorbax SB C18 (4.6 mm x 250 mm, 5 microm) column; mobile phase (A) water mobile phase (B) acetonitrile, gradient elution; UV detection wavelength 254 nm; ESI source and data acquisition in positive and negative mode.

RESULTThe accurate molecular weights of 20 compounds were measured and identified in ethyl acetate part of Huangqi Danggui decoction. Furthermore, the types of them are as below: flavonoids and flavonoid glycosides, saponins, oligosaccharides, amino acids and phthalides.

CONCLUSIONIt is a rapid and accurate method that the compositions of compound prescription of traditional Chinese medicine can be identified in terms of the separation of high performance liquid chromatography, the accurate molecular weights measured by MS and other information, which can clarify the potential effective compounds of Huangqi Danggui decoction.

Acetates ; chemistry ; Amino Acids ; chemistry ; Benzofurans ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; chemistry ; Flavonoids ; chemistry ; Glycosides ; chemistry ; Molecular Weight ; Oligosaccharides ; chemistry ; Saponins ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; methods

OBJECTIVETo study whether aristololactam I (AL-I) can enter renal proximal tubular epithelial cells and the situation of intracellular distribution and accumulation.

METHODCultured human renal proximal tubular epithelial cell line (HK-2) was used as the subject. Intracellular fluorescence from AL-I and its distribution are examined by fluorescence microscopy after a treatment with different concentration of AL-I, the intracellular accumulation of AL-I was also investigated by incubated cells in AL-I -free medium for 48 h after washing-out the media containing AL-I.

RESULTAfter treatment of AL-I (concentration from 5 microg x mL(-1) to 20 microg x mL(-1)), glaucous fluorescence could be observed inside renal proximal tubular epithelial cells at 0.5 h, and the fluorescence distributed only in cytoplasm while not be observed in nuclei. Moreover, the fluorescence of AL-I could be kept in cytoplasm for more than 48 h after washing out the media containing AL-I .

CONCLUSIONAL-I is able to enter renal proximal tubular epithelial cells in short time and accumulate in cytoplasm, but not enter nuclei. This property may contribute to the cytotoxic mechanism of renal injury induced by AL-I, which may partially explain the persistent renal toxicity of AAs and its metabolites in the development of aristolochic acid nephropathy.

Animals ; Aristolochic Acids ; metabolism ; toxicity ; Cell Line ; Cell Nucleus ; drug effects ; metabolism ; Cytoplasm ; drug effects ; metabolism ; Epithelial Cells ; cytology ; drug effects ; metabolism ; pathology ; Humans ; Kidney Diseases ; metabolism ; pathology ; Kidney Tubules, Proximal ; cytology ; pathology ; Microscopy, Fluorescence

OBJECTIVETo analyze and identify fatty acids in the seeds of Sterculia lychnophora.

METHODThe compositions was isolated and determined by GC-MS technique, and area normalization method was used to make quantitative analyze of the content of compositions.

RESULTS21 Fatty acids and 5 other compositions were isolated and determined.

CONCLUSIONThe major fatty acids are 9,12(Z,Z)-octadecadienoic acid(37.96%), hexadecanoic acid(24.77%), 9-(Z)-octadecenoic acid(19.77%) and octadecanoic acid(5.01%).

Fatty Acids, Nonesterified ; chemistry ; isolation & purification ; Fatty Acids, Unsaturated ; analysis ; Gas Chromatography-Mass Spectrometry ; Palmitic Acid ; analysis ; Plants, Medicinal ; chemistry ; Seeds ; chemistry ; Sterculia ; chemistry

OBJECTIVETo investigate whether age influence the precision of dual X-ray absorptiometry (DXA) measurement at the hip in middle-aged and elderly women.

METHODSTotally 90 women were randomly selected and divided into three age groups: 45-55 years, 56-65 years, and 66-75 years. Each age group contained 30 women. Each woman was scanned twice at the same day. Bone mineral density (BMD) values of femoral neck, ward's triangle, and trochanter were collected and grouped by calculating the root mean square (RMS). Precision errors were expressed as RMS (standard deviation, SD).

RESULTSFor the femoral neck and trochanter, significant differences of SD of BMD existed among all age groups. For the ward's triangle, significant difference of BMD existed among all age groups except between the 45-55 group and 56-65 age group.

CONCLUSIONAge can influence the precision of DXA measurement at the hip in middle-aged and elderly women.

Absorptiometry, Photon ; Age Factors ; Aged ; Analysis of Variance ; Bone Density ; Female ; Femur ; physiology ; Femur Neck ; physiology ; Humans ; Menopause ; physiology ; Middle Aged ; Reproducibility of Results

Construction of recombinant adenovirus, which contain human microdystrophin, and then transfection into mesenchymal cells( MSCs) of mdx mice were done, and genetically-corrected isogenic MSCs were acquired; the MSCs transplantation into the mdx mice was then done to treat the Duchenne muscular dystrophy( DMD). Microdystrophin cDNA was obtained from recombinant plasmid pBSK-MICRO digested with restrictive endonuclease Not I ; the production was inserted directionally into pShuttle-CMV. The plasmid of pShuttle-CMV-MICRO was digested by Pme I , the fragment containing microdystrophin was reclaimed and transfected into E. coli BJ5183 with plasmid pAdeasy-1. After screening by selected media, the extracted plasmid of positive bacteria was transfected into HEK293 cells with liposome and was identified by observing the CPE of cells and by the PCR method. Finally, MSCs of mdx mice were infected with the culture media containing recombinant adenovirus, and the expression of microdystrophin was detected by RT-PCR and immunocytochemistry. Recombinant adenovirus including microdystrophin was constructed successfully and the titer of recombinant adenovirus was about 5.58 x 10(12) vp/mL. The recombinant adenovirus could infect MSC of mdx mice and microdystrophin could be expressed in the MSC of mdx mice. Recombinant adenovirus including microdystrophin was constructed successfully, and the microdystrophin was expressed in the MSC of mdx mice. This lays the foundation for the further study of microdystrophin as a target gene to correct the dystrophin-defected MSC for stem cell transplantation to cure DMD.
Adenoviridae ; genetics ; Animals ; Cells, Cultured ; Dystrophin ; genetics ; metabolism ; Gene Expression ; Genetic Therapy ; methods ; Genetic Vectors ; genetics ; Humans ; Immunohistochemistry ; Mesenchymal Stem Cell Transplantation ; methods ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Mice ; Mice, Inbred mdx ; Muscular Dystrophy, Duchenne ; genetics ; pathology ; therapy ; Recombinant Fusion Proteins ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Transduction, Genetic

OBJECTIVETo determine the impact of Ureaplasma urealyticum (Uu) infection on the integrity of sperm plasma membrane in infertile males.

METHODSSixty-three semen samples were divided into a Uu infection group (n = 32) and a normal control group (n = 31). Conventional semen analyses were performed by computer-assisted semen analysis (CASA) and Uu detected by the culture method. The semen samples were washed with PBS and dyed by SYBR-14/PI double fluorescent staining, followed by detection of the integrity of sperm plasma membrane by flow cytometry. The percentage of the sperm with intact plasma membrane was indicated as the percentage of sperm emitting green fluorescence (SYBR-14+/PI-%).

RESULTSThe Uu infection group showed a significantly decreased integrity of sperm plasma membrane ([45.14 +/- 10.69]%) and reduced percentage of grade a + b sperm ([23.29 +/- 8.81]%) as compared with the normal control group ([72.68 +/- 9.91]% and [46.32 +/- 9.54]%) (P < 0.01). But there were no significant differences in the semen volume, pH value, and sperm concentration between the two groups (P > 0.05).

CONCLUSIONUu infection decreases the integrity of sperm plasma membrane, which might be an important factor of male infertility.

Adult ; Case-Control Studies ; Cell Membrane ; pathology ; Flow Cytometry ; Humans ; Infertility, Male ; microbiology ; pathology ; physiopathology ; Male ; Organic Chemicals ; Semen Analysis ; methods ; Spermatozoa ; metabolism ; pathology ; Ureaplasma Infections ; pathology ; physiopathology ; Ureaplasma urealyticum ; Young Adult

OBJECTIVETo evaluate the reliability of different hepatitis E diagnosis reagent tests on the acute hepatitis E.

METHODSThree acute hepatitis E diagnosis tests, E2-IgM (Wantai, China), GL-IgM and GL-IgG (Genelabs, Singapore) were compared for their reliability in a sera panel composed by 273 healthy individuals and 525 hepatitis.

RESULTSThe specificity of E2-IgM on the diagnosis of acute hepatitis E was 100.0%, it was significantly higher than GL-IgM (96.7%) and GL-IgG (85.4%). The sensitivity of E2-IgM and GL-IgG were 97.9% and 93.8% respectively, both significantly higher than GL-IgM (72.9%). Among 65 acute hepatitis cases being positive on GL-IgM test but negative on E2-IgM, 58 (89.2%) cases were found to be positive with anti-hepatitis A virus IgM, it indicated that the GL-IgM test might be interfered by other IgM antibodies on serum.

CONCLUSIONE2-IgM is a good test for the diagnosis of acute hepatitis E.

Acute Disease ; Hepatitis Antibodies ; blood ; Hepatitis E ; diagnosis ; Humans ; Immunoglobulin G ; blood ; Immunoglobulin M ; blood ; Reagent Kits, Diagnostic ; Sensitivity and Specificity

OBJECTIVETo develop an HPLC method for simultaneous determination of swertiamarin, gentiopicroside, sweroside, mangiferin, erythrocentaurin, and to detect these five constituents in eight Qingyedans derived from Swertia mileensis, S. cincta, S. patens, S. punicea, S. delavayi, S. nervosa, S. macrosperma and S. yunnanensis.

METHODThe separation was carried out on a Thermo BDS Hypersil C18 (4. 6 mm x 250 mm, 5 microm) column eluted with mobile phase of water containing 0. 1% phosphoric acid and methanol (B) in gradient program (0-10 min, 18%-20% B; 10-30 min, 20%-35% B; 30-35 min, 35%-60% B). The column temperature was 32 degrees C , and the detection wavelength was set at 250, 260, 225 nm. The flow rate was 0. 7 mL . min-1 from 0 to 30 min, and be increased to 1. 0 mL . min-1 in 35 min.

RESULTThe five compounds were well separated. The linear response ranges of swertiamarin, gentiopicroside, sweroside, mangiferin, erythrocentaurin were 0. 072-13. 39, 0. 1204. 518, 0. 060-5. 050, 0. 025-1. 518, and 0. 031-0. 210 microg, respectively. The mean recoveries of five compounds were 97.03% -102. 7% (RSD 1. 8% -6.2% ). There are swertiamarin, gentiopicroside and sweroside in most samples, and mangiferin in half samples. But erythrocentaurin was only detected in a few samples. The contents of five compounds were different in different samples. The contents of swertiamarin in S. mileensis, S. patens, S. yunnanensis and S. delavayi are up to 34. 47-118.05 mg . g-1, the contents of gentiopicroside are up to 25. 91 mg . g-1 in S. cincta. In S. puncea all contents of swertiamarin, gentiopicroside, sweroside and mangiferin are higher, especially the content of sweroside. There are Xiao-Qingyedans and Da-Qingyedans called in markets, and they can be identified by the contents of swertiamarin, gentiopicroside and sweroside. S. punicea can be identified by the content of sweroside, and the ratio gentiopicroside/total content can be used for identification of S. cincta from other seven Qingyedan species.

CONCLUSIONThe method was certified to be accurate and reliable and can be used for identification and quality evaluation of traditional Chinese medicine Qingyedan derived from Swertia species.

Chromatography, High Pressure Liquid ; methods ; Iridoid Glucosides ; analysis ; Pyrones ; analysis ; Swertia ; chemistry

OBJECTIVETo analyze the significant clinicopathologic factors related to tumor recurrence in patients with sinonasal squamous cell carcinomas (SCC) and to evaluate the effectiveness and plausibility of surgical salvage in the recurrent cases.

METHODSThe clinicopatholgic data of 107 patients with primary sinonasal SCC treated from 1996 to 2007 were analyzed retrospectively. Univariate and multivariate logistic regression analyses were used to define the risk factors related to tumor recurrence. Salvage surgery with was selectively carried out in the recurrent sinonasal SCC using different surgical approaches, including lateral rhinotomy midfacial degloving or combined craniofacial approach. Immediate reconstruction of major surgical defects were performed with latissimus dorsi flap, pectoralis major myocutanneous flap, temperalis fasciomuscular flap, free rectus abdominis flap and free radial forearm flap. All patients were routinely follwed up and 5-year survival were calculated using directly calculating method and Kaplan-Meier's method.

RESULTSThe 5-year survival rate of 107 cases was 52.3% (56/107). Local recurrence was the most common pattern of tumor relapse. Forty-four of the 107 cases had recurrence. Logistic regression analysis showed the T stag was the most important impacting factor for tumor recurrence (OR = 0.258, P = 0.001). Of 44 cases with recurrence, 33 cases underwent salvage surgery and the 5-year survival rate after salvage surgery was 29.1%.

CONCLUSIONST stag is the most important impacting factor for tumor recurrence. Salvage surgery with immediate reconstruction of major surgical defects should be carried out in the selective cases of recurrent sinonasal SCC.

Adult ; Aged ; Carcinoma, Squamous Cell ; pathology ; surgery ; Female ; Humans ; Male ; Middle Aged ; Neoplasm Recurrence, Local ; etiology ; surgery ; Reconstructive Surgical Procedures ; Retrospective Studies ; Salvage Therapy ; Survival Rate ; Young Adult