菟丝子指纹图谱研究新进展

本研究旨在建立菟丝子药材的高效液相色谱（HPLC）指纹图谱．为科学评价及有效控制菟丝子药材质量提供可靠方法。采用高效液相色谱法,以乙醇提取加水提取作为提取方法制备样品。分析了10批不同采摘时间的菟丝子药材,建立了菟丝子药材的HPLC指纹图谱,确立了10个共有峰,并且利用“中药色谱指纹图谱相似度评价软件”对不同样品之间的相似度进行计算。结果表明该方法准确可靠．重复性好,可为菟丝子药材的质量控制提供科学准确的依据。     

不同品种玫瑰花药材的HPLC指纹图谱研究

目的:建立中药玫瑰花的高效液相色谱指纹图谱,为科学评价和有效控制其质量提供可靠的方法。方法:用HPLC-UV方法测定了11份玫瑰花样品,采用中国药典委员会颁布的"中药色谱指纹图谱相似度评价"2004A系统建立了玫瑰花药材指纹图谱,并进行相似度评价和聚类分析。结果:11份玫瑰花样品的HPLC指纹图谱有18个共有峰,有10份样品相似度都在0.8以上,聚类分析时归为一类。结论:建立了玫瑰花不同品种的HPLC指纹图谱,并比较了其之间的差异,该方法稳定、可靠、精密度高、重复性好,可为中药玫瑰花的质量评价体系的建立提供科学依据。     

叶下珠药材HPLC指纹图谱的比较研究

采用反相高效液相色谱法,对叶下珠药材不同产地、不同采收期、不同部位指纹图谱进行测定和比较分析,并与中成药叶下珠胶囊的HPLC指纹图谱进行了比较研究。结果显示:(1)建立了叶下珠药材的HPLC特征指纹图谱,标定18个共有峰,利用对照品指认4个峰;11批(来源地不同)叶下珠样品的HPLC图谱相似度(相合系数,均值)在0.89~0.99之间。(2)7批不同采收期的叶下珠药材的HPLC指纹图谱相似度在0.94以上,各共有峰的峰面积大多随生长期而增加,至10月5日达到最高,建议叶下珠药材应于每年的10月上旬进行采收。(3)不同部位叶下珠药材HPLC指纹图谱相似度分析发现,叶、果的相似度较高(0.98~0.99),根、茎的相似度较低(0.86~0.87),说明根、茎中各成分含量较低,建议采收叶下珠药材的地上部分即可。(4)比较叶下珠胶囊和叶下珠药材的指纹图谱,发现二者的化学成分非常相似,但峰面积差异较大,其差异可能是由于加工过程所致。该研究所建立的HPLC指纹图谱分析方法简便、重现性好,可用于叶下珠药材及中成药的鉴定与质量评价。     

新疆雪莲的高效液相指纹图谱及液-质联用分析

建立新疆雪莲药材的高效液相指纹图谱。以Luna C18为分析柱,用乙腈-0.1%醋酸梯度洗脱,获得分离度较好的新疆雪莲药材HPLC分析条件。通过10批样品的分析和对照,标示出12个共有峰,相似度分析大于0.92。最后通过HPLC-UV-MS/MS联用分析鉴定出其中7个化学成分。该方法可为新疆雪莲药材指纹图谱和质量分析方法的建立提供参考。     

头花蓼和头状蓼的HPLC指纹图谱鉴定研究

建立头花蓼、头状蓼HPLC指纹图谱鉴定方法。用RP—HPLC（DAD）法,梯度洗脱,测定不同产地头花蓼及头状蓼的指纹图谱,并作相似度比较分析。不同产地头花蓼指纹图谱相似度较好,而头花蓼与头状蓼的指纹图谱有明显差异。HPLC指纹图谱具有重现性好,特征性强,方法简便等特点,可用于头花蓼、头状蓼药材的鉴别。     

广西铁皮石斛HPLC指纹图谱研究

采用HPLC法建立广西铁皮石斛提取液的指纹图谱,运用相似度评价药材质量。结果表明,在13个共有峰构成的铁皮石斛指纹图谱中,有11批药材相似度均在0.8以上,2批的相似度较差,未达到0.8,表明相似度大小与药材的产地有关。通过对栽培铁皮石斛不同部位药材的指纹图谱峰面积的比较,不提倡对药材提早采收。3批组织培养铁皮石斛的图谱相似度较低,说明培养到不同阶段的铁皮石斛材料组织培养物,其化学成分相差较大。     

不同产地土荆芥挥发油GC指纹图谱研究

采用水蒸气蒸馏法提取了12个产地土荆芥的挥发油,通过GC法分析其成分.运用中药色谱指纹图谱相似度建立共有模式,以相关度评价图谱的相似性,以共有峰与对照峰的比值对各产地土荆芥药材挥发油进行聚类分析.结果表明:12个产地的土荆芥药材的挥发油气相色谱指纹图谱有17个共有峰,其中气相色谱指纹图谱的相似度在0.85以上的有11个,12个产地的土荆芥药材的挥发油可通过系统聚类归为两类,不同产地土荆芥药材的挥发油相似度较高.该方法简便,所得挥发油中各成分分离度高,建立的指纹图谱重现性好,可作为土荆芥药材挥发油的质量控制方法之一.     

狼毒大戟二萜类成分HPLC指纹图谱及聚类分析

为建立狼毒大戟药材的特征图谱,科学评价其质量,本文采用高效液相色谱法,乙腈-水梯度洗脱,测定了11批不同来源的狼毒大戟样品二萜类化合物,通过相似度分析建立了狼毒大戟提取物的高效液相指纹图谱,并对11批药材进行系统聚类分析。本文采用乙腈-水为流动相,梯度洗脱,检测波长:230 nm,流速:1.0mL/min,进样量:10μL,柱温:30℃。相似度评价结果显示11批不同产地的狼毒大戟相似度均高于0.9,并对11批样品建立共有模式,确定了23个特征共有峰,聚类分析结果将狼毒大戟样品分为2类,与药材产地相关。本文所建立狼毒大戟的指纹图谱方法简便、可靠,可作为狼毒大戟药材的基源鉴定参考方法。     

不同产地野生与栽培伊贝母药材水溶性成分指纹图谱研究

目的:建立伊贝母药材水溶性成分高效液相色谱指纹图谱,为科学评价和有效控制其质量提供可靠的方法。方法:采用HPLC-DAD方法,以Kromasil C18色谱柱(4.6 mm×150 mm,5μm),流动相为甲醇和水,梯度洗脱,流速0.5 mL·min-1,柱温25℃,检测波长:260 nm。采用药典委员会颁布的中药色谱指纹图谱相似度评价系统2004 A版软件,对29批野生与栽培伊贝母药材进行指纹图谱分析。结果:29批伊贝母药材中有14个共有特征峰,建立了HPLC指纹图谱共有模式。各批次伊贝母药材相似度都在0.714以上,29批野生与栽培伊贝母药材可通过系统聚类分成5类,不同产地野生与栽培药材组成质量相似性较好,并定量测定了样品中的β-胸苷和腺苷。结论:所建立的HPLC指纹图谱及定量分析均具有良好的精密度、重复性、稳定性,可用于伊贝母药材的质量综合评价。     

基于GC-MS分析不同基原车前草挥发油成分的异同CSCD

比较不同基原车前草挥发油成分的异同。采用水蒸气蒸馏法提取14批次车前草挥发油,气相色谱-质谱联用技术(GC-MS)分离鉴定各色谱峰的化学成分;将样品挥发油指纹图谱导入指纹图谱相似度评价系统,以平车前草P1为参照图谱,进行多点校正,得相似度;采用聚类分析(HCA)、主成分分析(PCA)及正交偏最小二乘法-判别分析(OPLS-DA)对14批次样品挥发油检测结果进行分析。结果显示,从14批样品挥发油中共鉴定出62种化学成分,9种共有成分,均以棕榈酸、亚油酸、α-亚麻酸为主要成分;14批次样品指纹图谱高度相似,相似度在0.967~0.999;HCA和PCA均不能将两种不同基原的车前草按照品种分为两类。上述结果表明车前草与平车前草挥发油成分相似度较高,其作为同一药材入药具有一定的科学依据。     

Relationships between mitochondrial content of muscle fibres and patterns of glyogendepletion postmortem.

Sternomandibularis muscles were removed from slaughtered adult cattle immediately after exsanguination. On the basis of the density of diformazan granules deposited by a reaction for NAD tetrazolium reductase, approximately equal numbers of muscle fibres with high and low mitochondrial content were identified in serial frozen sections. In samples taken immediately after exanguination both types of muscle fibres exhibited glycogen phosphorylase activity and were stained equally by the periodic acid-Schiff (PAS) reaction for glycogen. In unstimulated muscle samples 1 hr postmortem, no loss of PAS staining was detected. In electrically stimulated samples 1 hr postmortem, large numbers of muscle fibres with a low mitochondrial content but only some muscle fibres with a high mitochondrial content became PAS-negative. Stimulation-induced glycogen depletion was completely prevented by the interfaicular injection of magnesium sulphate solution. In unstimulated samples between 5 and 24 hr postmortem, some muscle fibres with a high mitochondrial content but only a few muscle fibres with a low mitochondrial content became PAS-negative.     

Application of enzyme field-effect transistors for determination of glucose concentrations in blood serum

Glucose-sensitive enzyme field effect transistors (ENFETs) modified by an additional Nafion membrane have been developed and used for diluted blood samples analysis. The ENFET was used in the linear portion of the calibration curve up to 1.5 mM glucose in a model solution, which corresponds with up to 60 mM glucose in the undiluted samples (dilution 1:40). The high linearity of the Grans curve (factor of linearity is 1.03) obtained by the method of standard additions indicates the high precision of analysis. Glucose concentrations in different blood serum samples determined by ENFETs were compared with those measured by the commercial analyzer 'Eksan-G' and colorimetric method ('Diagluc' enzymatic kit), and good correlation between these methods was revealed. The high reproducibility and operational stability of the biosensor developed were demonstrated.     

PCR—微孔板杂交法检测不同人群TTV DNA

根据报道的TTV全序列设计引物和探针,建立PCR－微孔板杂交法,检测81例正常人群、92例职业献血员123例甲－庚型肝炎、32例非甲－庚型肝炎、48型发性肝癌患者的TTV DNA。结果表明TTV在以上五种人群中的阳性率分别为3．7％、4．3％、21．1％、28．1％、52．0％,前者与后三者比较有显著性差异（P＜0．05）,TTV合并HBV二重感染重叠感染的54．0％,这揭示不同人群均存在TTV感染,正常人群和职业献血员存在健康携带状态,甲－庚型肝炎和非甲－庚肝炎病人为高危人群,TTV可与各型肝炎存在重叠感染,TTV除经血传播外,存在其它传播途径,TTV感染与ALT及TBIL的升高密切相关。     

High expression of MDM2 protein and low rate of p21(WAF1/CIP1) expression in SCID mice Epstein Barr virus-induced lymphoproliferation.

To study the prevalence of p53 inactivation and MDM2/p21(WAFI/CIP1) expression in severe combined immunodeficient (SCID) mice Epstein-Barr virus (EBV)-induced lymphoproliferation, 19 samples obtained after ip injection of peripheral blood mononuclear cells (PBMCs) from EBV-seropositive donors or lymphoblastoid cell lines (LCL) were analyzed. In all samples tested, overexpression of Ki-67 antigen was shown by immunohistochemistry, indicating a high proliferative index of SCID mice EBV-induced lymphoproliferation. P53 mutations were screened by functional assay in yeast in 14 samples. With this test, a p53-inactivating mutation was found in only one case; the remaining cases exhibited a wild-type p53 pattern. However, an accumulation of p53 protein was detected by immunohistochemistry in six of 19 samples. P21 expression was found in seven of 19 samples but was not correlated with the rate of p53 protein in tumors. In contrast, high levels of nuclear accumulation of MDM2 were found in all samples by immunohistochemistry. These results suggest that a high Ki-67 proliferative index in SCID mice EBV-induced lymphoproliferation is not due to the inactivation of p53 by mutation, but could be associated with an overexpression of MDM2, which would act by a p53-independent mechanism.(J Histochem Cytochem 47:1315-1321, 1999)     

环介导等温扩增技术在食品中痢疾志贺氏菌快速检测

【目的】探讨、优化基于环介导等温扩增技术(LAMP)快速检测常规食品中感染性痢疾志贺氏菌的方法。【方法】在NCBI数据库中搜索获取志贺氏菌的特异性基因高度保守区,设计3对LAMP反应引物,建立、优化该LAMP可视化检测方法,并评价其特异性、灵敏度,同时与PCR检测方法和传统检测方法对比,进行结果统计分析。【结果】5株志贺氏菌标准菌株样品均检测为阳性,11株非志贺氏菌标准菌株样品均检测为阴性,无交叉反应。最低检验限为1.6×101 CFU/反应(或1.6×101 CFU/m L),且经比较,LAMP检测灵敏度比PCR检测高出1个数量级。通过对161份实际样品和人工污染样品进行检测,LAMP检测与传统方法检测结果具有较高的一致性。【结论】LAMP具有检测过程快速简便、检测结果稳定、可靠的特点,适用于对常规食品中感染性痢疾志贺氏菌的高效、快速检测需求。     

Monitoring occupational exposure to carcinogens: detection by 32P-postlabelling of aromatic DNA adducts in white blood cells from iron foundry workers

Blood samples were volunteered by workers in a Finnish iron foundry who were occupationally exposed to polycyclic aromatic hydrocarbons and from control subjects not known to be occupationally exposed to this class of chemical carcinogens. DNA was isolated from peripheral white blood cells and digested with micrococcal nuclease, spleen phosphodiesterase and nuclease P1. The DNA digest was then incubated with [gamma-32P]ATP and polynucleotide kinase. Aromatic adducts present in the digest that were resistant to nuclease P1 were thus 32P-labelled while unmodified nucleotides were not. The 32P-labelled adducts were resolved by t.l.c. and detected by autoradiography. Foundry workers were classified as belonging to high, medium or low exposure groups according to their exposure to airborne benzo[a]pyrene (high greater than 0.2, medium 0.05-0.2, low less than 0.05 microgram BP/m3 air). Aromatic adducts were found to be present in DNA from 3/4 samples from the high exposure group, 8/10 samples from the medium exposure group. 4/18 samples from the low exposure group and 1/9 samples from the unexposed controls. The levels of adducts found in the high and medium group samples ranged up to 1 adduct in 10(7) nucleotides but the levels formed in the low exposure group samples were not significantly different from those in unexposed controls. No differences related to the smoking habits of the subjects were observed. Most of the DNA adducts detected had chromatographic mobilities distinct from those formed when the 7,8-diol 9,10-oxide of BP reacted with DNA. The results indicate that highly-exposed individuals are more likely to contain aromatic DNA adducts in their white blood cells, but large interindividual variations were evident. In addition, multiple samples from the same subjects indicate that qualitative and quantitative changes in adduct patterns occur with time. This pilot study suggests that 32P-postlabelling may be useful in monitoring human exposure to known and to previously unidentified environmental genotoxic agents.     

Effects of sulfate concentration on the anaerobic dechlorination of polychlorinated biphenyls in estuarine sediments

In order to determine the effects of sulfate concentration on the anaerobic dechlorination of polychlorinated biphenyls, sediments spiked with Aroclor 1242 were made into slurries using media which had various sulfate concentrations ranging from 3 to 23 mM. The time course of dechlorination clearly demonstrated that dechlorination was inhibited at high concentration of sulfate due to less dechlorination of meta-substituted congeners. When the dechlorination patterns were analyzed by the calculation of Euclidean distance, the dechlorination pathway in the 3 mM sulfate samples was found to be different from that observed in the 13 mM samples, although the extent of dechlorination in these two samples was similar. It is possible that the dechlorination in the high sulfate concentration samples is inhibited by the suppression of growth of methanogen, which have been shown to be meta-dechlorinating microorganisms.     

Carbohydrate components and crystalline structure of organosolv hemp (Cannabis sativa L.) bast fibers pulp

Changes in carbohydrate components and crystalline structure of hemp bast fibers during organosolv pulping were investigated by X-ray diffractometry, FT-IR spectroscopy and high performance liquid chromatography (HPLC). The reasons for defibrillation and beating problems with organosolv hemp bast fiber pulp were investigated with reference to these properties of pulp samples. Hemp bast fibers and organosolv pulp samples had low hemicellulose contents and high cellulose contents. It was found that the disorder parameter of cellulose in hemp bast fibers was very low, when crystalline cellulose ratio was high and the crystalline structure of cellulose in hemp bast fibers was very stable. These properties affected defibrillation and beating of organosolv hemp bast fibers pulp negatively.     

Releasing of plasminogen-activator from the kidney to the blood (author's transl)]

When fibrinolytic activity in blood samples from various vessels was examined by the dilute-blood-clotlysis-time method (DBCLT), it was found to be noticeably high in the renal venous blood, though the activity was not detected by usual blood clotlysis time method. Plasmin was not detected in any blood samples examined, and the contents of fibrinogen and fibrin (or fibrinogen) breakdown products in the renal venous blood were not significantly different from those in the blood from other vessels. However, the high activity of plasminogen-activator was found only in the renal venous blood. Inhibitors on plasmin and plasminogen-activator (urokinase) were detected in almost the same amount in the blood samples from the various vessels. The amount of the inhibitors was sufficient to inhibit the plasminogen activation by urokinase, whose activity was equivalent to the plasminogen-activator activity in the renal venous blood. These results indicate that the high activity by DBCLT in the renal venous blood was derived from the high activity of plasminogen-activator, which was inactivated by inhibitors in undiluted blood. Plasminogen-activator may be released from the kidney to the blood, and immediately inactivated by the inhibitors in renal vein, and then diluted with systemic blood which contains little plasminogen-activator.     

An A-ELISA to detect hepatitis A virus in estuarine samples.

An amplified enzyme-linked immunosorbent assay (A-ELISA) for detecting and quantifying hepatitis A virus in estuarine water samples is described. The test was five times more sensitive than a standard ELISA and at least two times more sensitive than radioimmunoassay. Test sensitivity was unaffected by the procedures used to concentrate the virus in estuarine samples or by the presence of humic and tannic acids in test samples. Nonspecific reactions were not encountered with a number of enteroviruses or with a rotavirus. A high sensitivity and specificity combined with speed, low cost, and freedom from radiolabels made the A-ELISA useful for detecting hepatitis A virus in environmental samples. The virus was detected in 3 of 20 estuarine water samples examined by A-ELISA.