高效液相色谱法测定兔眼房水中环孢霉素A的含量

目的建立房水中环孢霉素A的高效液相色谱测定方法,并测定兔用药后眼房水中的药物浓度。方法采用WatersC18色谱柱,流动相为乙腈-甲醇-水(66∶25∶20),流速为0.8ml/min,在55℃柱温下,于214nm处检测。结果本方法CsA线性范围10~150ng/ml,最低检测浓度10ng/ml,回收率99.81%,日间和日内RSD均小于8%。结论本法具有简便、灵敏、快速、稳定性好、柱温低的特点,适合用于眼房水中CsA浓度监测。     

Distribution of azithromycin into brain tissue, cerebrospinal fluid, and aqueous humor of the eye.

To measure the concentrations of azithromycin in the central nervous system, 20 patients with brain tumors (group I) received a single 500-mg oral dose of azithromycin either 24, 48, 72, or 96 h prior to the tumor removal operation and 10 patients with cataracts undergoing surgery (group II) and 7 patients scheduled to undergo lumbar puncture (group III) received the same dose of azithromycin 24 h prior to the operation or procedure. Serum from all patients, brain tissue from group I, aqueous humor from group II, and cerebrospinal fluid from group III were assayed for azithromycin concentration. The mean concentrations of azithromycin in brain tissue 24, 48, 72, and 96 h after administration were 2.63 +/- 2.58, 3.64 +/- 3.81, 0.74 +/- 0.37, and 0.41 micrograms/g, respectively. In contrast, the concentrations of azithromycin in cerebrospinal fluid and aqueous humor of the eye were very low or undetectable. Therefore, these data show that azithromycin appears to be widely distributed into brain tissue but not into cerebrospinal fluid or aqueous humor of the eye.     

毫米波辐射对兔眼房水一氧化氮及某些细胞因子的影响

目的观察毫米波辐射后兔眼房水一氧化氮（ＮＯ）及白细胞介素６（ＩＬ６）、白细胞介素８（ＩＬ８）、肿瘤坏死因子（ＴＮＦα）含量的变化,探讨毫米波对眼的损伤机制,为制定毫米波辐射的卫生标准提供实验依据。方法新西兰兔按受辐照后不同取材时间及辐射剂量分组,采用３５ＧＨｚ等幅连续波,辐射兔眼１ｈ,用硝酸酶还原法测定房水ＮＯ,用ＥＬＩＳＡ法测定ＩＬ６、ＩＬ８、ＴＮＦα。结果１０ｍＷ／ｃｍ２毫米波辐射结束后６ｈ,房水ＮＯ含量升高（Ｐ＜０．０１）,此效应可持续２４ｈ以上;３５ｍＷ／ｃｍ２及１０ｍＷ／ｃｍ２毫米波辐射结束后２４ｈ内,未发现房水ＩＬ６、ＩＬ８、ＴＮＦα含量的升高。结论低功率３５ＧＨｚ毫米波辐射可引起房水中ＮＯ含量的增高,提示ＮＯ为毫米波致眼损伤的重要因素之一     

角膜穿通损伤合并海水浸泡模型免角膜组织病理学及房水白细胞介素6的变化

背景：由于海水特殊的理化性质,海上创伤后细胞的病理改变常有特殊性。目的：观察兔角膜爆炸穿通伤合并海水浸泡角膜组织病理学改变及房水白细胞介素6水平变化。方法：取成年健康灰兔16只,单支鞭炮致兔角膜爆炸伤,于角膜中周部做长3mm全层切口。右眼为实验侧,左眼为对照侧。实验侧将海水通过角膜切口注入前房,海水持续冲洗眼表30min。对照侧使用生理盐水。结果与结论：光镜观察显示实验侧造模后1、2和3d角膜上皮细胞大片坏死脱落,基质层水肿明显伴大量炎细胞浸润。对照侧1、2d角膜病理改变同实验组,但程度较轻,造模后3、5d局部病理改变明显减轻。实验组造模后第1、2、3d房水中自细胞介素6水平明显高于对照侧（尸〈0．05）。表明海水组伤情较对照组明显加重,提示海水可能是重要的致伤因素之一。     

Determination of phosphatidylglycerol in amniotic fluid by a simple one-dimensional thin-layer chromatography method

Phosphatidylglycerol (PG) in amniotic fluid is the second important component of lung surfactant phospholipids and may be clinically useful in assessing fetal lung maturity in utero. Although methods for PG determination are available, there are shortcomings in clinical application. We developed an alternative reliable onedimensional thin-layer chromatography (TLC) method for separating and quantitating PG in amniotic fluid. A mini-TLC plate (8 × 10 cm) was prepared from silica gel H containing 5% ammonium sulfate. The plate was first developed in tetrahydrofuron/dimethoxymethane/methanol/2N ammonium hydroxide (30.0:20.6:5.6:3.0, v/v) and then in chloroform/methanol (60:9, v/v) in the same direction. PG was clearly separated from other phospholipids and neutral lipids, even when large amounts of other phospholipids were present on the TLC plate. The density of the charred PG was directly proportional to the amount of PG up to 8 nmol. The content of PG in nmol in the specimen can be quantitated by comparing with a standard PG. Up to 10% of blood serum or 3% meconium showed no detectable PG, nor did these substances affect PG quantitation in amniotic fluid. This method is sensitive and accurate. It is also time-saving and economical.     

Effects of travoprost eye drops on intraocular pressure and pulsatile ocular blood flow: a 180-day, randomized, double-masked comparison with latanoprost eye drops in patients with open-angle glaucoma

Background: Because of the increasing realization of the importance of optic nerve head perfusion in the pathogenesis of glaucoma, the influence of new antiglaucomatous drugs on ocular hemodynamic properties should be investigated.Objective: The aim of this study was to compare the effects of 2 prostaglandin analogues, travoprost eye drops and latanoprost eye drops, on intraocular pressure (IOP) and pulsatile ocular blood flow (pOBF) in patients with primary open-angle glaucoma (POAG).Methods: Previously untreated patients aged 40 to 60 years with POAG and normal brachial blood pressure (BBP), heart rate, body mass index, and hemorheologic findings were eligible for this randomized, double-masked study. Two drops of travoprost (group T) or latanoprost (group L) were self-administered in both eyes at 9:00 pm. In all patients, IOP, pOBF, BBP, and heart rate were measured at baseline and on days 15, 30, 60, 90, and 180 of treatment.Results: Twenty-five consecutive patients with POAG were enrolled in this study conducted at the Glaucoma Research Center of the Department of Ophthalmology, Bari University, Policlinico di Bari (Bari, Italy). Of these, 7 were withdrawn because they did not return for the second appointment, leaving 18 patients (11 men, 7 women; mean [SD] age, 51.9 [5.5] years) to complete the study. In both groups, mean IOP values were significantly reduced at all time points compared with baseline (all P<0.01). Mean pOBF values increased ∼50% from baseline following treatment with either travoprost or latanoprost by day 15, were maintained at that level for 60 days, and then gradually decreased (group T: P = NS, NS, <0.01, <0.05, and <0.05 at days 15, 30, 60, 90, and 180, respectively, vs baseline; group L: P<0.01 at all time points vs baseline). All other parameters remained constant throughout the study. An early inverse correlation between IOP and pOBF was noted in group T but not in group L. No significant differences were found between groups in IOP or pOBF at any time point.Conclusions: In this study population, pOBF was increased with travoprost and latanoprost in the short term, but this effect was kept constant only with travoprost. IOP was reduced with both drugs after short-term therapy, and this reduction was maintained in both groups. Travoprost may represent another option for the medical treatment of POAG.     

角膜穿通损伤合并海水浸泡模型兔角膜组织病理学及房水白细胞介素6的变化(英文)

背景:由于海水特殊的理化性质,海上创伤后细胞的病理改变常有特殊性。目的:观察兔角膜爆炸穿通伤合并海水浸泡角膜组织病理学改变及房水白细胞介素6水平变化。方法:取成年健康灰兔16只,单支鞭炮致兔角膜爆炸伤,于角膜中周部做长3mm全层切口。右眼为实验侧,左眼为对照侧。实验侧将海水通过角膜切口注入前房,海水持续冲洗眼表30min。对照侧使用生理盐水。结果与结论:光镜观察显示实验侧造模后1、2和3d角膜上皮细胞大片坏死脱落,基质层水肿明显伴大量炎细胞浸润。对照侧1、2d角膜病理改变同实验组,但程度较轻,造模后3、5d局部病理改变明显减轻。实验组造模后第1、2、3d房水中白细胞介素6水平明显高于对照侧(P<0.05)。表明海水组伤情较对照组明显加重,提示海水可能是重要的致伤因素之一。     

密度梯度离心与贴壁法分离培养免骨髓间充质干细胞及其生物学特性观察

背景：获取更大量、更高纯度、有活性的骨髓间充质干细胞是干细胞移植及组织工程研究发展的基础。目的：拟建立一套体外分离培养及大量扩增兔骨髓间充质干细胞的方法，并对细胞生物学特性进行观察。设计、时间及地点：细胞学体外观察，于2006—12／2007—07在山东省烟台市莱阳中心医院骨科试验室完成。材料：清洁级1月龄雌性新西兰大白兔1只。方法：采用密度梯度离心法和贴壁培养法相结合的方式，分离兔骨髓间充质干细胞，应用细胞刮收集呈长梭形的细胞，形成克隆后用胰蛋白酶＋EDTA消化，按1：3传代进行体外扩增培养。主要观察指标：倒置相差显微镜下观察细胞形态特征，免疫细胞化学SABC法鉴定CD34、CD44抗原的表达，观察细胞生长特性，测定细胞活力。结果：原代分离培养的贴壁细胞呈长梭形，漩涡状排列，约14d达90％以上融合；传代后增殖迅速，细胞为单一的梭形，排列更加有序。所培养的细胞CD44呈阳性表达，而CD34呈阴性。第1～5代细胞培养3—5d为对数生长期，第7代以后对数增长期延长。第1～5代细胞成活率均〉94％，其中第3，4代成活率高达97％；至第7代细胞活力呈明显下降趋势，细胞成活率仅为78％。结论：应用密度梯度离心结合贴壁法，可成功分离并扩增兔骨髓间充质干细胞，且细胞生长稳定，增殖力强．可作为细胞移植的种子细胞。     

密度梯度离心与贴壁法分离培养兔骨髓间充质干细胞及其生物学特性观察

背景：获取更大量、更高纯度、有活性的骨髓间充质干细胞是干细胞移植及组织工程研究发展的基础。目的：拟建立一套体外分离培养及大量扩增兔骨髓间充质干细胞的方法，并对细胞生物学特性进行观察。设计、时间及地点：细胞学体外观察，于2006—12／2007—07在山东省烟台市莱阳中心医院骨科试验室完成。材料：清洁级1月龄雌性新西兰大白兔1只。方法：采用密度梯度离心法和贴壁培养法相结合的方式，分离兔骨髓间充质干细胞，应用细胞刮收集呈长梭形的细胞，形成克隆后用胰蛋白酶＋EDTA消化，按1：3传代进行体外扩增培养。主要观察指标：倒置相差显微镜下观察细胞形态特征，免疫细胞化学SABC法鉴定CD34、CD44抗原的表达，观察细胞生长特性，测定细胞活力。结果：原代分离培养的贴壁细胞呈长梭形，漩涡状排列，约14d达90％以上融合；传代后增殖迅速，细胞为单一的梭形，排列更加有序。所培养的细胞CD44呈阳性表达，而CD34呈阴性。第1～5代细胞培养3—5d为对数生长期，第7代以后对数增长期延长。第1～5代细胞成活率均〉94％，其中第3，4代成活率高达97％；至第7代细胞活力呈明显下降趋势，细胞成活率仅为78％。结论：应用密度梯度离心结合贴壁法，可成功分离并扩增兔骨髓间充质干细胞，且细胞生长稳定，增殖力强．可作为细胞移植的种子细胞。     

对比多种眼药治疗先天性上睑下垂术后相关性干眼症的临床疗效

目的比较妥布霉素眼膏单独使用及联合小牛血去蛋白提取物眼用凝胶、聚乙二醇滴眼液治疗先天性上睑下垂术后相关性干眼症的临床疗效。方法采用回顾性研究,对2012年8月~2016年5月于重庆医科大学附属儿童医院眼科诊断为先天性上睑下垂并接受额肌瓣悬吊术的术后住院患儿478例(640只眼)的临床资料进行统计分析。根据上睑下垂程度分为轻、中、重度3类,根据术后单独使用妥布霉素眼膏(A组),妥布霉素联合小牛血去蛋白提取物眼用凝胶(B组),妥布霉素联合聚乙二醇滴眼液分为三组(C组)。分别统计三组患儿术后第3天的症状评分、角膜荧光素染色评分(FL)、泪膜破裂试验(BUT)及基础泪液分泌试验(Sit)。结果对比A组,B组在轻、中、重3种不同程度的上睑下垂术后干眼有显著疗效(P0.05);而C组在轻、中程度的上睑下垂术后的干眼有明显疗效(P0.05);B与C组进行比较,显示前者在治疗中、重度上睑下垂术后疗效更好(P0.05)。结论相较于单独使用妥布霉素眼膏,额肌瓣悬吊术后使用妥布霉素眼膏联合小牛血去蛋白提取物眼用凝胶或聚乙二醇滴眼液对术后并发的干眼症具有明显优越性,且联合使用小牛血去蛋白提取物眼用凝胶更利于减轻干眼症症状及预防暴露性角膜炎。     

兔角膜碱烧伤模型房水中肿瘤坏死因子α及血管内皮生长因子表达与姜黄素的抑制

背景：稳定的角膜新生血管动物模型是研究角膜新生血管调控机制,姜黄素对碱烧伤角膜新生血管具有抑制作用和保护作用。目的：探讨姜黄素对碱烧伤角膜新生血管模型中肿瘤坏死因子α及血管内皮生长因子表达的影响,为防治角膜新生血管提供理论依据。方法：纳入33只新西兰大耳白兔,随机取3只设为正常组,其余30只建立兔角膜碱烧伤诱发角膜新生血管模型,右眼设为对照组给予生理盐水,左眼设为干预组给予姜黄素,裂隙灯观察角膜新生血管生长及角膜混浊情况,酶联免疫吸附实验检测肿瘤坏死因子α和血管内皮生长因子在房水中的表达。结果：正常组没有角膜新生血管生成。与对照组比较,干预组角膜新生血管受到抑制且角膜混浊较轻（P〈0.05）。房水肿瘤坏死因子α和血管内皮生长因子在3组中均有表达,对照组和干预组明显高于正常组,但干预组低于对照组（P〈0.05）。说明姜黄素可以有效降低角膜碱烧伤后房水肿瘤坏死因子α及血管内皮生长因子的表达进而抑制兔角膜碱烧伤后角膜新生血管的生长。     

Determination of polyamines and metabolites in cerebrospinal fluid by isotope dilution mass fragmentography, and a clinical application

Capillary gas chromatography and mass fragmentography was used to determine simultaneously 1,3-diaminopropane, putrescine, cadaverine, spermidine, spermine, isoputreanine and putreanine in cerebrospinal fluid. After addition of deuterium labelled analogs and acid hydrolysis, the compounds were isolated by adsorption onto silica and converted into their N-heptafluorobutyryl-methylesters. Quality control data and an application of the method are given. A patient harbouring an astrocytoma was monitored during chemotherapeutic treatment.     

Determination of lysozyme in serum, urine, cerebrospinal fluid and feces by enzyme immunoassay

Conjugates of human lysozyme and horseradish peroxidase (HRP) were prepared by means of the heterobifunctional reagent N-succinimidyl 3-(2-pyridyldithio) propionate. A conjugate containing 2 mol HRP/mol lysozyme was isolated by gel filtration and used as a labeled antigen in competitive enzyme immunoassays, in which anti-lysozyme rabbit IgG had been bound to wells of microtiter plates. The assay can detect as little as 1 microgram lysozyme/l. The following reference intervals have been established: 950-2450 micrograms/l for serum, 1.7-123 micrograms/l for urine, 17.6-118 micrograms/l for cerebrospinal fluid and 0.04-1.5 microgram/g for feces.     

Determination of flumequine and a hydroxy metabolite in biological fluids by high-pressure liquid chromatographic, fluorometric, and microbiological methods

A sensitive and specific high-pressure liquid chromatographic method is described for the determination of the antibacterial drug flumequine and a major metabolite, 7-hydroxyflumequine, in human plasma and urine. The assay was linear over a concentration range of 1 to 120 micrograms/ml for both compounds. This method is compared with fluorometric and microbiological assays for flumequine. These latter methods did not differentiate between flumequine and any fluorescent or antimicrobiologically active metabolites. However, because essentially all drug in the plasma was found to be flumequine in radiolabeled studies, levels of unchanged drug in the plasma could be quantitated by either high-pressure liquid chromatography or fluorometry. Although only high-pressure liquid chromatography was able to specifically measure flumequine in the urine, the antimicrobial activity of the urine, which is more therapeutically relevant due to antimicrobially active metabolites, could be quantitated by either the fluorometric or the microbiological assay.     

Determination of lysozyme in biological fluids by a semi-automatic kinetic technique. Discussion of the method (author's transl)]

A rapid and semi-automatic determination of lysozyme in biological fluids using kinetic analysis and turbidimetry is described and compared to other commonly used techniques. The specificity of the method is satisfactory while that of clarification of a gel medium is apparently not. Normal values and standard errors for plasma, urine, and leucocytes are given. A standardised expression of lysozyme activity is proposed and discussed. The importance of the means by which the blood sample is collected and prepared is underlined: plasma, decanted soon after collection, is preferable to serum.     

Rapid, sensitive, and validated method for detection of Salmonella in food by an enrichment broth culture - nested PCR combination assay

A rapid nested PCR assay for detection of Salmonella from food was developed. The sensitivity of the assay developed was comparable to the traditional culture based methods with an advantage in reduction of assay time. The assay procedure with artificially contaminated samples was able to detect as low as 4CFU Salmonella/25g of food samples (sprout, carrot, cucumber and poultry meat). With two synthetic primers of 26 mer TS11 and 25 mer TS4, a 1.2kb fragment was amplified which served as a template for amplification of final 375bp product using TS11 and TS5 primers. No non-specific amplification from the native microbial flora of food samples was observed. The reaction generates a single band specific to Salmonella which allows the analyst to interpret data at ease and without any confusion. Enriched broth serves as template for the reaction which removes labour intensive DNA isolation procedures. In case of artificially contaminated samples, 6h enriched lactose broth can serve as template. However, for market samples where the organisms are under environmental stress, it is desirable to use template from Rappaport Vasiliadis medium. The assay also employes internal amplification control, which is amplified into a 300bp fragment and thus serves as positive control for the reaction and any possibility of false negative due to inhibitory action of food components on PCR reaction can be ruled out.     

Determination of creatinine in serum and urine by a rapid liquid-chromatographic method

We describe an HPLC ion-pair procedure for rapid and specific evaluation of creatinine in serum and urine. We used a 15 cm X 4.6 mm ODS column with a 50/50 (by vol) mixture of sodium decanesulfonic acid (10 mmol/L, pH 3.2) and methanol and measured absorbance at 236 nm. Serum (100 microL) or 30-fold-diluted urine (100 microL) was added to 400 microL of acetone. After centrifugation, the supernates (300 microL) were dried, reconstituted with the mobile phase, and injected into the HPLC. Assay precision was tested for concentrations of 10, 29, and 130 mg/L and yielded, respectively, 3.1%, 2.1%, and 1.1% for within-day CV and 2.8%, 2.1%, and 2.2% for total CV. Analytical recovery was 102 (+/- 6.7%). Linearity was demonstrated in the 0-200 mg/L range for serum and 0-3.5 g/L range for urine (r greater than or equal to 0.999). The detection limit for creatinine (signal-to-noise ratio = 3) was 0.5 mg/L. We used cimetidine for internal standardization. Correlation was good between this procedure and the Jaffé kinetic, the enzymatic (creatinine amidohydrolase), and the Fuller's earth alkaline picrate methods.