四逆汤煎煮过程中乌头类生物碱的溶出和水解平衡

目的 研究四逆汤中乌头类生物碱的溶出平衡和水解平衡。方法 利用电喷雾质谱分析，比较了制附子、四逆汤、四逆汤药渣和含有3种双酯型生物碱的混合对照品体系中的乌头碱类二萜生物碱。结果 在煎煮过程中双酯型生物碱溶解并发生水解反应，而脂类生物碱则难溶于水。乌头碱、中乌头碱、次乌头碱水溶性相近，但是次乌头碱在水中的热稳定性更高，C19二萜骨架上C3取代基(—OH或—H)的变化影响生物碱的稳定性。结论四逆汤中的乌头碱类生物碱的种类及含量由其溶解性和化学稳定性共同决定。     

八味地黄汤各药味对附子生物碱影响的电喷雾质谱研究

目的以八味地黄汤为研究对象,考察各药味对附子生物碱的影响规律,揭示附子配伍组方的科学内涵,为科学阐释八味地黄汤配伍规律提供化学依据。方法利用电喷雾质谱技术分别对八味地黄汤,附子加山茱萸、山药、肉桂、丹皮、泽泻、地黄、茯苓共煎液以及次乌头碱加山茱萸、山药共煎液进行系统考察。结果次乌头碱加山药、山茱萸,附子加山药等共煎液中次乌头碱含量较高;八味地黄汤、附子加山茱萸共煎液中次乌头碱含量较低;肉桂、丹皮、泽泻、地黄、茯苓等对附子生物碱的影响不明显。结论八味地黄汤中乌头碱和中乌头碱水解和溶出比较完全,次乌头碱的含量较少,次乌头碱的减少是由于水解以及溶出受到抑制导致。其中山茱萸对制附子起减毒作用,其减毒机制主要是抑制了制附子中次乌头碱的溶出。而山药则能导致附子毒性增加。     

复方盐酸二甲双胍片在人体内药物动力学和生物等效性研究

目的 建立人血浆中盐酸二甲双胍的反相离子对高效液相色谱和格列本脲的液相色谱-质谱测定方法,研究复方盐酸二甲双胍片(盐酸二甲双胍250mg/格列本脲1.25mg×2片)相对于联合使用的盐酸二甲双胍片(500mg)和格列本脲片(2.5mg)在男性健康志愿者体内的药物动力学行为,评价其生物利用度和生物等效性。方法 采用双交叉随机实验设计:20名受试者交叉口服复方盐酸二甲双胍片2片或口服盐酸二甲双胍片与格列本脲片各1片,服药后于0.5、1.0、1.5、2.0、2.5、3.0、3.5、4.0、5.0、6.0、8.0、12、24、36h分别取血,分离血浆,分别依法测定盐酸二甲双胍和格列本脲的血药浓度。结果 测得口服复方盐酸二甲双胍片或联合使用盐酸二甲双胍片与格列本脲片后,盐酸二甲双胍的达峰时间分别为(2.0±0.7)h和(2.1±0.9)h,峰浓度分别为(1402.4±349.2)μg·L^-1和(1329.7±315.4)μg·L^-1,消除半衰期分别为(3.84±0.61)h和(4.26±0.96)h,AUC_0-24分别为(7292.7±1967.5)μg·L^-1和(7416.2±1843.9)μg·h·L^-1；格列本脲的达峰时间分别为(3.1±0.9)h 和(3.0±0.8)h，峰浓度(71.7±22.7)μg·L^-1和(70.3±20.7)μg·L^-1,消除半衰期分别为(5.05±2.01)h 和(4.78±1.64)h，AUC_0-24分别为(367.6±168.7)μg·L^-1和(352.6±144.7)μg·h·L^-1；以联合服用等剂量的盐酸二甲双胍片与格列本脲片为参比，以AUC_0-24计算得复方盐酸二甲双胍片之盐酸二甲双胍和格列本脲的相对生物利用度，分别为101.1%±28.5%和123.7%±82.9%。结论 建立的分析方法准确灵敏，测得数据可靠。统计学分析表明复方盐酸二甲双胍片与联合使用盐酸二甲双胍片和格列本脲片显生物等效。     

芫花药材的HPLC指纹图谱及ESI-MS分析

目的:建立芫花药材的质量评价方法。方法:采用HPLC-DAD对21批芫花药材的指纹图谱进行比较研究,并用HPLC-ESI-MS技术对部分共有峰进行了初步归属。结果:建立了以16个共有峰为特征指纹信息的芫花药材的HPLC指纹图谱,并通过MS技术初步鉴定出其中10个共有峰所对应的可能化学成分,共有峰主要为黄酮苷及其苷元类成分。结论:该方法可为芫花药材指纹图谱和质量分析方法的建立提供参考。     

LC／ESI—MS测定小鼠血浆中异烟肼及其代谢物

目的:建立测定小鼠血浆中异烟肼及其代谢物乙酰肼、肼的液相色谱-电喷雾质谱联用(LC/ESI—MS)法。方法:血浆样品经甲醇沉淀蛋白后,用衍生化试剂3-甲氧基苯甲醛 MBA 进行柱前衍生化,衍生化产物以甲醇-5 mmol·L~(-1)醋酸铵(pH5.0)缓冲溶液(60:40)为流动相,采用 Agilent Zorbax XDB—C_8柱(5 μm,4.6 mm×150 mm)分离,通过 ESI~ -MS 以选择离子监测(SIM)模式进行检测,用于定量分析的离子分别为 m/z 256.1(异烟肼衍生物),m/z 193.2(乙酰肼衍生物),m/z 151.1(肼衍生物)。结果:异烟肼、乙酰肼、肼的线性范围分别为1～64,0.05～3.2,0.025～1.6μg·mL~(-1);定量下限分别为1,0.05,0.025μg·mL~(-1);方法的平均回收率分别为98.9%,98.0%,97.2%;平均 RSD 值分别为2.7%,2.4%,3.1%。结论:本法简便、准确,适用于小鼠血浆中异烟肼、乙酰肼和肼血药浓度的测定。     

Characterization of Mixtures of Alkyl Polyglycosides (Plantacare) by Liquid Chromatography-Electrospray Ionization Quadrupole Time-of-Flight Mass Spectrometry

No HeadingPurpose. Industrial products of alkyl polyglycosides (APGs) are widely used as well tolerated surfactants in washing and cleaning agents. APGs belong to the class of nonionic surfactants and are mixtures of molecules consisting of a hydrocarbon chain with Y carbon atoms linked to X sugar residues. Physicochemical properties of APG products strongly depend on the molecular composition. Therefore, a detailed analytical investigation of technical-grade APGs is presented.Methods. Complex mixtures of alkyl/alkenyl polyglycosides (APGs, Plantacare) and mixtures of APGs with other surfactants were analyzed by reversed-phase liquid chromatography-electrospray ionization quadrupole time-of-flight mass spectrometry (LC/ESI-QTOF-MS) using methanol-water as mobile phase and gradient elution.Results. Analytes were separated according to the chain length of the alkyl homologs. Under the soft ionization of the ESI technique, mainly [M+Na]⁺ ions were observed, which proved to be very stable. Additionally [M+H]⁺ ions were detected. The QTOF mass spectrometer allows to identify even high molecular mass components in the mixtures, and APGs up to eight sugar residues were found. From these data, the alkyl/alkenyl chain lengths and the degree of oligomerization of the sugar moiety for different technical-grade APGs were calculated. Using MS/MS experiments, additional structural and chemical information was obtained.Conclusions. The presented LC/ESI-QTOF-MS approach allows to analyze and characterize various APG products, such as Plantacare. The ability of this LC/ESI-QTOF-MS approach to analyze mixtures of APGs with other surfactants is demonstrated.     

内蒙古赤芍化学成分的HPLC—DAD／ESI—MS分析

目的:利用液-质联用技术定性分析内蒙古赤芍药材中的化学成分。方法:以甲醇回流提取内蒙古赤芍药材。高效液相色谱条件:色谱柱为 Zorbax sB-C~(18)柱(4.6 mm×250 mm,5 μm),流动相0.2%的醋酸水溶液(A)-0.2%醋酸乙腈溶液(B),梯度洗脱,流速0.5 mL·min~(-1),柱温30℃,二极管阵列检测器(DAD),检测波长230 nm。质谱条件:负离子模式扫描,扫描范围100～2000 amu;氮气流速10 L·min~(-1),雾化温度350 ℃,毛细管电压3500 V,裂解电压100 V。结果:通过高效液相色谱将赤芍化学成分较好地分离,根据紫外光谱可大致判断其化合物类型,由电喷雾质谱得到各成分的相对分子质量,进而推测出其中13个主要成分的可能结构。结论:液-质联用技术能快速定性研究赤芍药材中的化学成分,为赤芍指纹图谱的优化和质量标准的制定提供依据。     

白眉蝮蛇蛇毒糖基化蛋白的质谱分析

[目的]蛋白质组学方法分析和初步鉴定旅顺产白眉蝮蛇（Gloydius blomhoffi Brevicaudus）蛇毒中的糖蛋白。[方法]SDS-PAGE用于白眉蝮蛇粗毒蛋白质组份分离,糖蛋白凝胶荧光染料Pro-Q-Emerald染糖基化蛋白,采用胰蛋白酶酶解蛋白质,用反相C18-HPLC分离肽段混合物,用纳升电喷雾电离串联质谱（nESI-MS/MS）系统进行的质谱数据采集,蛋白质鉴定用Biowroks软件搜索NCBInr数据库完成。[结果]旅顺产白眉蝮蛇蛇毒L-氨基酸氧化酶、金属蛋白酶、谷氨酰环化酶、salmobin、纤溶酶原激活物、contortrixobin、磷脂酶A2,神经增长因子和较小分子量的L-氨基酸氧化酶被初步鉴定为糖蛋白,其匹配的肽段数分别为8、4、7、2、3、3、4、2和5个。L-氨基酸氧化酶的纯化与表征结果验证了其糖基化,并以多种糖基化形式存在。[结论]采用蛋白质组学手段从旅顺产白眉蝮蛇蛇毒中鉴定出了8种糖蛋白,该方法简便、快速、准确。     

猴头菌寡糖的化学研究

 目的研究猴头菌寡糖的理化性质、组成糖、相对分子质量及糖链连接方式,并确定猴头菌四糖的2种化学结构。方法采用水提醇沉法;醇沉部分经DEAE-Sephadex A-50和Sephadex G-25分离纯化;苯酚硫酸法、Lowry法、间羟基联苯法测定总糖、蛋白质、糖醛酸的含量;HPLC测定纯度及相对分子质量分布;活性碳柱色谱纯化猴头菌四糖;GC,GC-MS及ESI-MS分别给出组成糖、糖链连接方式及化学结构。结果Hep-1-3为寡糖混合物,总糖含量为91%,蛋白质含量为3%,糖醛酸含量甚微,由Man和Glu组成,糖苷键以1→4和1→6为主,3-位上有支点。所得2种四糖均由Glu组成,以1→6连接为主链,3-位上有支点。结论Hep-1-3为二糖至七糖的寡糖混合物,首次从猴头菌中获得,所得2种四糖的结构确定为a-D-Glul→6a- D-Glul→6a-D-Glul→6a-D-Glu和a-D-Glul→6a-D-Glul→6a-D-Glu↓₁³ a-D-Glu     

Toxicity and detoxification of lipid-derived aldehydes in cultured retinal pigmented epithelial cells

Age-related macular degeneration (ARMD) is the leading cause of blindness in the developed world and yet its pathogenesis remains poorly understood. Retina has high levels of polyunsaturated fatty acids (PUFAs) and functions under conditions of oxidative stress. To investigate whether peroxidative products of PUFAs induce apoptosis in retinal pigmented epithelial (RPE) cells and possibly contribute to ARMD, human retinal pigmented epithelial cells (ARPE-19) were exposed to micromolar concentrations of H2O2, 4-hydroxynonenal (HNE) and 4-hydroxyhexenal (HHE). A concentration- and time-dependent increase in H2O2-, HNE-, and HHE-induced apoptosis was observed when monitored by quantifying DNA fragmentation as determined by ELISA, flow cytometry, and Hoechst staining. The broad-spectrum inhibitor of apoptosis Z-VAD inhibited apoptosis. Treatment of RPE cells with a thionein peptide prior to exposure to H2O2 or HNE reduced the formation of protein-HNE adducts as well as alteration in mitochondrial membrane potential and apoptosis. Using 3H-HNE, various metabolic pathways to detoxify HNE by ARPE-19 cells were studied. The metabolites were separated by HPLC and characterized by ElectroSpray Ionization-Mass Spectrometry (ESI-MS) and gas chromatography-MS. Three main metabolic routes of HNE detoxification were detected: (1) conjugation with glutathione (GSH) to form GS-HNE, catalyzed by glutathione-S-transferase (GST), (2) reduction of GS-HNE catalyzed by aldose reductase, and (3) oxidation of HNE catalyzed by aldehyde dehydrogenase (ALDH). Preventing HNE formation by a combined strategy of antioxidants, scavenging HNE by thionein peptide, and inhibiting apoptosis by caspase inhibitors may offer a potential therapy to limit retinal degeneration in ARMD.