HPLC法测定白果中银杏酸的含量

目的:建立反相高效液相色谱法测定银杏白果中有毒成分银杏酸含量。方法:用LC／DAD／ESI／MS对白果中的银杏酸进行定性鉴定。白果经石油醚索氏提取、浓缩、定容后,直接进样进行HPLC测定。色谱分析条件:色谱柱为Inertsil ODS-2(250 mm×4.6 mm,5μm),流动相为甲醇-3％HAc溶液(92:8),流速1.0 mL·min-1,柱温40℃,紫外检测波长310 nm。结果:白果中含有5种银杏酸:C15:1,C17:2,C15:0和C17:1(存在2种同分异构体),还有1种作者推测为C17:3,其中C17:1和推测为C17:3的银杏酸为首次在白果中发现。该法精密度和重复性较好,其5次测定的RSD分别为0.71％和3.7％。白果中银杏酸含量达200 mg·kg-1左右,大大超过国际限量标准,其食用安全性应引起高度重视。结论:该法简便、快速、准确,适用于白果及其产品中银杏酸的分析,可为白果产品的质量监控和安全性评价提供基础。     

一种新的抗革兰阳性菌制剂达托霉素

介绍一种新的脂肽类抗生素———达托霉素的特点和临床应用 ,达托霉素在体外和体内的抗菌活性及副作用。体外 ,达托霉素具有快速、浓度依赖性的杀菌活性和相对延长的抗生素后效应 ;体内 ,达托霉素呈线性药物动力学 ,每日 1次的给药累积量小。提示 :达托霉素具有抗所有革兰阳性菌的作用 ,包括一些对传统耐药的细菌 ,其副作用轻微、安全性高 ,是一种有应用前途的抗生素。     

白桦酸类化合物的研究进展

白桦酸类化合物是一类可以从多种植物中分离得到的天然产物，包括白桦酸、23-羟基白桦酸、白桦素及其衍生物等，是一类结构及作用机制相对新颖、具有抗HIV-1和抗肿增活性的天然化合物。对这类化合物的结构修饰和改造，目前已经获得大量具有良好生物活性的衍生物。现对这方面的研究进展进行综述。     

UPLC/MS比较分析达托霉素和内酯水解物的酸降解产物

摘要：达托霉素是一种正在临床使用的新型抗耐药菌抗生素。但是由于其结构特点，达托霉素容易受酸碱影响产生降解。 本文利用UPLC/MS比较分析了达托霉素及其内酯水解物酸降解后的产物，发现达托霉素主要的降解产物为天冬氨酰-丙氨酰内 酰胺键断裂产物，内酯水解物酸的主要降解产物为分子量974的七肽和分子量1160的九肽。通过比较，确定分子量974的有关物 质主要来源于内酯水解物的酸降解。并且通过核磁共振检测，确定了该七肽物质的化学结构和氨基酸连接顺序。     

高效液相色谱法测定银杏叶提取物及其制剂中银杏酸的含量高效液相色谱法测定银杏叶提取物及其制剂中银杏酸的含量

目的建立银杏叶提取物及其制剂中银杏酸含量的HPLC测定方法。方法银杏叶提取物及其制剂经石油醚提取，提取液浓缩后用石油醚定容，HPLC直接测定，并用LC/MS对其中的银杏酸进行了定性鉴定。色谱分析条件：色谱柱为Inertsil ODS-2，流动相为甲醇-3% HAc溶液（92∶8），流速1.0 mL·min^-1,柱温40 ℃，紫外检测波长310 nm。结果银杏叶提取物中存在6种银杏酸C13:0，C15:0，C15:1，C17:1，C17:2和一种可能是C17∶3的银杏酸化合物，其中C13∶0，C15∶1和C17∶1占总银杏酸的94%以上。实验测得高银杏酸含量的提取物中银杏酸含量为1.12%,RSD为2.4%（N=5）；银杏叶提取物片剂中银杏酸含量为49.2 μg·g^-1,RSD为4.3%（N=5）。平均回收率98.2%，RSD为2.6%（N=5）。结论该方法准确、快速、简便，可用于银杏叶提取物及其制剂中银杏酸的定量分析。     

达托霉素可致嗜酸粒细胞性肺炎

2010年7月29日,FDA发布安全信息：静脉注射用达托霉素（Daptomycin）可能引起嗜酸粒细胞性肺炎。达托霉素为环酯肽类抗生素,用于金黄色葡萄球菌菌血症,复杂性皮肤和皮肤软组织感染。达托霉素的作用机制与其他抗生素不同,它通过扰乱细胞膜对氨基酸的转运、阻碍细菌细胞壁肽聚糖和胞壁酸酯的生物合成、改变细胞膜电位等多种机制破坏细胞膜功能,使其内容物外泄而达到杀菌的目的㈨。2004至2010年,FDA收到7份使用达托霉素与发生嗜酸粒细胞性肺炎有关的病例报告,其中6例来自FDA的不良事件报告系统,1例来自医学文献。     

高效液相色谱法测定银杏叶提取物及其制剂中银杏酸的含量

目的建立银杏叶提取物及其制剂中银杏酸含量的HPLC测定方法.方法银杏叶提取物及其制剂经石油醚提取,提取液浓缩后用石油醚定容,HPLC直接测定,并用LC/MS对其中的银杏酸进行了定性鉴定.色谱分析条件:色谱柱为Inertsil ODS-2,流动相为甲醇-3% HAc溶液(92∶8),流速1.0 mL*min-1,柱温40 ℃,紫外检测波长310 nm.结果银杏叶提取物中存在6种银杏酸C13:0,C15:0,C15:1,C17:1,C17:2和一种可能是C17∶3的银杏酸化合物,其中C13∶0,C15∶1和C17∶1占总银杏酸的94%以上.实验测得高银杏酸含量的提取物中银杏酸含量为1.12%,RSD为2.4%(n=5);银杏叶提取物片剂中银杏酸含量为49.2 μg*g-1,RSD为4.3%(n=5).平均回收率98.2%,RSD为2.6%(n=5).结论该方法准确、快速、简便,可用于银杏叶提取物及其制剂中银杏酸的定量分析.     

氧化还原状态的改变控制手霉素诱导HL-60白血病细胞凋亡

目的研究手霉素对HL-60白血病细胞的作用,探讨其作用机制,主要是线粒体的改变。方法使用人的白血病细胞株HL-60细胞。MTT细胞毒性测定评价对白血病细胞的作用,使用Annexin V和一氧化氮(nitric oxide,NO)染料标记细胞,应用流式细胞仪术检测细胞内NO生成和细胞凋亡。细胞内超氧阴离子通过二氢乙啶(dihydroethidium,DHE)测定。应用化学发光法测定超氧歧化酶(superoxide dismutase,SOD)活性。采用荧光法测定谷胱甘肽(glutathi-one,GSH),萤光素-萤光素酶发光法测定ATP含量。免疫印迹技术检测细胞色素C和Mn-SOD的表达。结果手霉素引起HL-60细胞活性的下降,且呈剂量依赖性。手霉素诱导产生反应性氧基(reactive oxygen species,ROS):NO和超氧阴离子,降低GSH,但不影响SOD。手霉素诱导线粒体降低细胞ATP的含量,线粒体肿胀和细胞色素C从线粒体释放到细胞质。手霉素诱导的凋亡与ROS的增加有关。用N-乙酰基-L-半胱氨酸(N-acetyl-L-cysteine,NAC)抑制ROS可保护HL-60细胞逃避手霉素的细胞毒作用和避免手霉素诱导的凋亡。结论细胞内ROS的产生对手霉素的细胞毒作用起非常重要的作用。手霉素通过包括上游ROS产生,线粒体形态改变和细胞色素C释放的线粒体途径,诱导白血病细胞凋亡。     

银杏酸的高效液相色谱法测定

目的建立银杏酸的简便预净化处理和HPLC含量测定方法。方法用LC/DAD/ESI/MS对银杏酸进行定性鉴定。银杏叶经正己烷提取、硅胶柱色谱净化处理,HPLC测定。色谱分析条件:色谱柱为Inertsil ODS-2,流动相为甲醇-3% HAc溶液(92∶8),流速1.0 mL·min^-1,柱温40℃,紫外检测波长310 nm。结果银杏叶中存在5种银杏酸C13∶0,C15∶0,C15∶1,C17∶1和C17∶2,其中C15∶1和C17∶1约占银杏酸的85%,C17∶2银杏酸未见国内文献报道。银杏叶提取物经硅胶柱色谱处理后,其HPLC谱中除银杏酸峰外,几无其他杂质峰。平均回收率97.0%,RSD为1.7%(N=6)。结论该方法准确、简便、可靠,可用于银杏酸的定量分析。     

白桦酸及其衍生物的研究进展

对白桦酸衍生物的构效关系及其中的YK-FH312、RPR103611和IC9564等的作用机制及活性进行述评。天然产物白桦酸是有抗HIV和抗肿瘤活性，且具有新型结构和新型作用机制的化合物，其天然来源广泛，半合成方法成熟，是寻找抗HIV和抗肿瘤药物的优秀的先导物。通过对白桦酸的结构修饰，大量白桦酸衍生物已被合成。     

Cytotoxicity of ginkgolic acid in HepG2 cells and primary rat hepatocytes

Ginkgolic acids and related alkylphenols (e.g. cardanols and cardols) have been recognized as hazardous compounds with suspected cytotoxic, allergenic, mutagenic and carcinogenic properties. To determine whether the phase I metabolism could contribute to their cytotoxicity, we investigated the cytotoxicity of one model compound, ginkgolic acid (15:1), using in vitro bioassay systems. In the first step, cytochrome P450 enzymes involved in ginkgolic acid metabolism were investigated in rat liver microsomes; then, two in vitro cell-based assay systems, primary rat hepatocytes and HepG2 cells, were used to study and the measurement of MTT reduction was used to assess cell viability. Results indicated that the cytotoxicity of ginkgolic acid in primary rat hepatocytes was lower than in HepG2 cells. Ginkgolic acid was demonstrated less cytotoxicity in four-day-cultured primary rat hepatocytes than in 20-h cultured ones. Co-incubation with selective CYP inhibitors, α-naphthoflavone and ketoconazole, could decrease the cytotoxicity of ginkgolic acid in primary rat hepatocytes. In agreement, pretreatment with selective CYP inducers, β-naphthoflavone and rifampin, could increase the cytotoxicity of ginkgolic acid in HepG2 cells. These findings suggest that HepG2 cells were more sensitive to the cytotoxicity of ginkgolic acid than primary rat hepatocytes, and CYP1A and CYP3A could metabolize ginkgolic acid to more toxic compounds.     

银杏酸的反相银化高效液相色谱法分离和测定

目的　建立银杏酸的反相银化高效液相色谱分离和含量测定方法。方法　用LC ESI MS对银杏叶中银杏酸进行鉴定,并用反相银化HPLC研究4种银杏酸的分离和含量测定。在银杏叶的乙醇浸提液中加入酸性盐溶液和少量吸附剂后,用正己烷萃取,N₂ 浓缩后的残留物进行HPLC分析。流动相为甲醇-5 %乙酸(90∶10 ) ,其中银离子浓度为0.03mol·L^-1 ,HPLC和二极管阵列检测联用对银杏酸色谱峰进行光谱分析和纯度鉴定。结果　反相银化HPLC分析银杏酸所确定的色谱峰是正确的,4种银杏酸与杂质成分均完全分离。平均回收率97.3% ,方法的精密度RSD为1.6 %。结论　反相银化HPLC是分离碳数与饱和度均不相同的同系物的有效方法,可用于银杏酸的定量分析和银杏叶生产的质量控制     

Non-specific SIRT inhibition as a mechanism for the cytotoxicity of ginkgolic acids and urushiols

Ginkgolic acids and urushiols are natural alkylphenols known for their mutagenic, carcinogenic and genotoxic potential. However, the mechanism of toxicity of these compounds has not been thoroughly elucidated so far. Considering that the SIRT inhibitory potential of anacardic acids has been hypothesized by in silico techniques, we herein demonstrated through both in vitro and computational methods that structurally related compounds such as ginkgolic acids and urushiols are able to modulate SIRT activity. Moreover, their SIRT inhibitory profile and cytotoxicity were comparable to sirtinol, a non-specific SIRT inhibitor (SIRT1 and SIRT2), and different from EX-527, a SIRT1 specific inhibitor. This is the first report on the SIRT inhibition of ginkgolic acids and urushiols. The results reported here are in line with previously observed effects on the induction of apoptosis by this class of compounds, and the non-specific SIRT inhibition is suggested as a new mechanism for their in vitro cytotoxicity.     

水杨酸-g-壳聚糖衍生物的合成及药效研究

制备水杨酸-g-壳聚糖载体衍生物，观察两者在药效上的协同和互补作用。通过二甲苯致小鼠耳肿胀实验观察水杨酸与壳聚糖抗炎的协同作用；通过小鼠扭体实验和热板痛实验观察衍生物的镇痛作用；通过胃黏膜形态学变化评价壳聚糖与水杨酸的互补作用。实验结果表明，水杨酸-g-壳聚糖衍生物外用抗炎作用优于水杨酸、壳聚糖和皮炎平，内服优于阿司匹林；即时镇痛作用低于阿司匹林，长效镇痛作用与阿司匹林相近；对胃黏膜的刺激性远远低于阿司匹林；水杨酸-g-壳聚糖衍生物具有抗炎协同作用和药效互补作用。     

Studies on the reactions between daptomycin and glyceraldehyde

The objectives of this project were to determine the reaction pathways of daptomycin in the presence of glyceraldehyde in acidic solutions, and to quantitate the kinetics of the major pathways. In the presence of glyceraldehyde (pH range 1-7 at 25 to 60 degrees C), daptomycin formed two major products separable by RP-HPLC. The products were identified using UV spectroscopy, fluorimetry, mass spectrometry, and 2D-1H NMR. The reaction scheme involved the reversible formation of imine and anilide derivatives. Carbinolamine was believed to be a common intermediate in formation pathways of both products. The carbinolamine intermediate underwent either acid catalyzed dehydration resulting in imine formation or intramolecular hydrogen bonding and bond cleavage giving rise to anilide formation. In mild acid conditions, both products reversed to daptomycin. The reaction between daptomycin and glyceraldehyde was first-order with respect to both reactants. In a pH range of 1-7, the imine formation rate was pH dependent with a maximum rate at approximate pH values of 3-4. The observed pH dependence was consistent with the pH dependence of typical amine-aldehyde reactions.     

Reactive oxygen species mediated membrane damage induced by fullerene derivatives and its possible biological implications

Fullerenes have attracted considerable attention in recent years due to their unique chemical structure and potential applications. Hence it is of interest to study their biological effects. Using rat liver microsomes as model systems we have examined the ability of the most commonly used fullerene, C60 and its water-soluble derivative, C60(OH)18 to induce membrane damage on photosensitization. For photoexcitation, UV or tungsten lamps were used. Damage was assessed as lipid peroxidation products like conjugated dienes, lipid hydroperoxides and thiobarbituric acid reactive substances (TBARS). protein oxidation in the form of protein carbonyls, besides loss of membrane bound enzymes. Both fullerene derivatives induced significant oxidative damage. The alterations induced were both time- and concentration-dependent. Role of different reactive oxygen species (ROS) in the damage induced was examined by various scavengers of ROS and by deuteration of the buffer. The changes induced by C60 were predominantly due to 1O2 while that by C60(OH)18 was mainly due to radical species. Biological antioxidants such as glutathione, ascorbic acid and alpha-tocopherol were capable of inhibiting membrane damage induced by both the fullerenes. However, the damage induced by C60(OH)18 was more for both lipids and proteins than that showed by C60. C60 also showed enhancement in the formation of lipid peroxidation in sarcoma 180 ascites microsomes. In conclusion, our studies indicate that fullerene/its derivative can generate ROS on photoexcitation and can induce significant lipid peroxidation/protein oxidation in membranes and these phenomena can be prevented by endogenous/natural antioxidants.     

Apoptotic effect in the glioma cells induced by specific protein extracted from Okinawa Habu (Trimeresurus flavoviridis) venom in relation to oxidative stress.

Okinawa Habu (Trimeresurus flavoviridis) venom is well known for its toxic efficacy, from which one kind of specific protein, Okinawa Habu apoxin protein-1 (OHAP-1) has been extracted. The purpose of this study was to investigate whether OHAP-1 could induce apoptosis in some glioma cells, and if so, to elucidate the possible mechanism involved. Three malignant glioma cell lines were tested. The malignant glioma cell lines were rat C6 and human RBR 17T, U251. OHAP-1 inhibited growth of all cell lines. Whether or not the apoptosis had been induced was determined by using DNA gel electrophoresis, DNA flow cytometry and TUNEL assay. After OHAP-1 treatment, DNA fragmentation, an increase in the percentage of subdiploid DNA content, and TUNEL positive cells were found in the C6, RBR17T, and U251 cells. Furthermore, OHAP-1 showed L-amino acid oxidase (LAAO) activity. In order to study the mechanism of apoptosis induced by OHAP-1, the changes of intracellular reactive oxygen species (ROS) were measured using flow cytometry, and the expression of p53 protein was examined using immunohistochemistry. OHAP-1 was found to generate ROS and increase the expression of p53 protein in glioma cells. The inhibiting effect of OHAP-1 on three tested cells was reversed when an antioxidant of either catalase or reduced glutathione (GSH) was added; its apoptotic effect correspondingly became weaker. In this study, the apoptotic effect of OHAP-1 on some malignant glioma cells was confirmed, and it could be that this effect might be mediated through promoting the generation of intracellular ROS and p53 protein expression in glioma cells. It was suggested that OHAP-1 is promising as a potential candidate for clinical tumor therapy.     

Radical scavenger activity of phenylethanoid glycosides in FMLP stimulated human polymorphonuclear leukocytes: structure-activity relationships

Radical scavenger activities of 21 phenylethanoid glycosides, including 15 ester derivatives of caffeic, ferulic, vanillic and syringic acid as well as 6 deacyl derivatives were determined by quantifying their effects on the production of reactive oxygen species (ROS) in a luminol-enhanced chemiluminescence assay with formyl-methionyl-leucyl-phenylalanine (FMLP) stimulated human polymorphonuclear neutrophils (PMNs). All phenylethanoids acylated with phenolic acids showed strong antioxidant activity whereas the deacyl derivatives were more than 30-fold less active. Therefore, the antioxidant activity is mainly related to the number of aromatic methoxy and hydroxy groups and the structure of the acyl moiety (C6-C1 or C6-C3). In contrast, modification of the sugar chain or replacement of hydroxy groups by methoxy groups in the acyl or the phenylethanoid moiety is of minor importance. The position of the acyl moiety is without significance. Free caffeic, ferulic, vanillic and syringic acid are less active compared to the phenylethanoid derivatives. This points to the importance of dissociation and lipophilicity of these acids in a cellular test system.