附子及乌头属植物中二萜类生物碱的电喷雾质谱研究

电喷雾质谱是一种软电离技术，生物碱分子中均含有氮原子，具有很高的质子亲合势和气相碱度，特别适合电喷雾质谱分析。本文利用电喷雾质谱系统研究了乌头属的几种植物中二菇类生物碱的质谱行为。并且对中药复方中的生物碱成分的化学变化进行探讨，在串联质谱中，乌头碱类生物碱的裂解方式相似。双酷型、三醋型、脂类生物碱在MS2中容易丢失C8乙酸或长链脂肪酸形成特征子离子峰，在进一步CID谱中，在C15位失去CO为其特征。但是单酷型生物碱在串联质谱中碎裂方式与上述几种生物碱有很大差别，例如，在MSZ中主要失去甲醇，还会失去C3位经基，但是不会失去CS轻基。利用这些规律分析了附子及草乌中的生物碱，发现了多种具有特殊取代基的新化合物。以电喷雾质谱为检测手段分析了附子在炮制过程中的成分变化；并且发现了分离提取脂类生物碱的方法。高乌头中的生物碱以C18型为主，同时也包含少量C19乌头碱骨架的生物碱。这两种类型的生物碱在串联质谱中的碎裂规律相似，都容易失去取代基，而生物碱骨架比较稳定，难以碎裂。但是不同位置的取代基失去时所形成的子离子的丰度有很大差别，经过串联质谱可以获取重要的结构信息。并且通过串联质谱确定了刺乌头碱环糊精的包合位点，更正了文献中的错误。关白附的CZ。型海替生型生物碱在串联质谱中的行为与上述生物碱相似，容易失去取代基，但是在失去取代基的同时，常常伴随骨架中C-C键的断裂。由于这些生物碱的结构存在较强的规律，所以很容易表征其中的结构相似的新生物碱。另一方面，我们发现导致白术与附子配伍将导致共煎液中有毒生物碱成分增加，从而解释了八味地黄丸与人参汤导致中毒的原因。并且通过对八味地黄汤的配伍研究，发现起解毒作用的药材为山茱英，解毒机理为抑制了有毒生物碱的溶出。方中的山药能够抑制双酷型生物碱的水解。

Apoptotic activity of frog Bombina maxima skin albumin

Albumin, the most abundant protein components of blood plasma, is synthesized and secreted by liver cells in vertebrates. Recently, it was demonstrated that frog Bombina maxima albumin is also expressed in skin. Both B. maxima albumins from skin and serum (BmA-skin and BmAserum) have similar biochemical characteristics except that the former contains haem b. Present studies showed that BmA-skin exhibited cytotoxic activity on H9 and C8166 cells. Pretreated with hemin to induce erythroid differentiation, K562 cells lost their resistance to cytotoxicity of BmAskin. After treating cells with BmA-skin for 48 h, 50 percentage cytotoxic concentrations (CC50) of BmA-skin on H9, C8166 and hemin-treated K562 cells were 1.31±0.09, 1.59±0.08 and 2.28±0.06 μM, respectively. The cell death induced by BmA-skin was mediated by apoptosis of the tested cell lines, as demonstrated by nuclear morphological changes, DNA fragmentation and DNA hypodiploidy of apoptosis cells. At BmA-skin concentration of 2 μM, 27.3%, 19.7% and 17.8% of H9, C8166 and hemin-treated K562 cells were found to be apoptotic. In contrast, BmA-serum possessed no cytotoxic and apoptosis-inducing activity on all the cell lines tested, even with concentration used up to 15 μM. These results indicated that bound haem b in BmA-skin contributed significantly to its cytotoxic and apoptosis-inducing activity on the cell lines assayed.

滇金足草、凋缨菊和长喙吴萸的化学成分研究

本论文对滇金足草（Goldfussia yunnanensis）、凋缨菊（Camchaya loloana）和长喙吴萸（Evodia vestia）的化学成分进行了研究，通过色谱分离得到40个化合物。主要基于波谱数据鉴定了它们的结构，其中10个为新化合物。 1．从滇金足草地上枝叶的95%乙醇提取物中共分离鉴定了16个化合物：泽漆内酯A（1）、18-羟基泽漆内酯A（2）、18-氧代泽漆内酯A（3）、18-羟基-3-O-β-D-吡喃葡萄糖-泽漆内酯A（4）、3-O-β-D-吡喃葡萄糖-泽漆内酯A（5）、3-O-β-D-吡喃半乳糖-泽漆内酯A（6）、6-E-肉桂酰哈巴俄苷（7）、E-哈巴俄苷（8）、5，6-异亚丙二氧基哈巴俄苷（9）、β-谷甾醇（10）、β-胡萝卜苷（11）、齐墩果酸（12）、肉桂酸（13）、麦角固醇（14）、硬脂酸（15）和丁二酸（16）。其中2-7为新化合物。5，6-异亚丙二氧基哈巴俄苷（9）以人工产物形式得到。 2．从凋缨菊地上枝叶的95%乙醇提取物中分离并鉴定了13个化合物：凋缨菊内酯A~C (17-19)、1β-乙酰基凋缨菊内酯C(20)、b-谷甾醇（10）、β-胡萝卜苷（11）、羽扇豆醇（21）、桦木醇（22）、桦木酸（23）、芥子醇（24）、紫丁香苷（25）、咖啡酸（26）和熊果酸（27）。其中化合物17-20为桉叶烷内酯类新化合物。化合物17、18、20对细胞株HepG2的GI50依次为7.80、7.08、4.99 µg/mL。 3．从长喙吴萸（E. vestia）地上枝叶的95%乙醇提取物中分离并鉴定了13个化合物：佛手内酯（28）、花椒毒素（29）、异茴芹内酯（30）、七叶内酯（31）、东莨宕素（32）、瑞香素（33）、异紫花前胡内酯(34)、茵芋碱（35）、山刈碱（36）、白鲜碱（37）、黄柏酮（38）、柠檬苦素（39）和对羟基苯甲醛（40）。 4．综述了1990—2007年期间从菊科植物中发现的桉叶烷-12，6内酯的化学结构、生物活性、生物转化及化学合成方面的研究进展。 Phytochemical investigation on Goldfussia yunnanensis, Camchaya loloana, and Evodia vestia, led to the isolation of 40 compounds, 10 of which were new ones. 1. Six new compounds were isolation from 95% ethanolic extract of the aerial parts of G. yunnanensis, and identified as 18-hydroxyhelioscopinolide A (2), 18-oxohelioscopinolide A (3), 18-hydroxy-3-O-β-D-glucopyranosylhelioscopinolide A (4), 3-O-β-D-glucopyranosylhelioscopinolide A (5),3-O-β-D-Galactopyranosyl helioscopinolide A (6), 6-O-trans-cinnamoyl E-harpagoside (7). The known compounds isolated were helioscopinolide A (1), E-harpagoside A (8), 5,6-isopropylidene E-harpagoside A (9), β-sitosterol (10), β-daucosterol (11), oleanolic acid (12), cinnamic acid (13), ergosterol (14), stearic acid (15) and succinic acid (16). Compound 9 was an artifact. 2. Four new compounds, loloanolides A – C (17 - 19) and 1β-acetoxy-loloanolide C (20), were isolation from 95% ethanolic extract of the aerial parts of C. loloana. The known ones were β-sitosterol (10), β-daucosterol (11), lupeol (21), betulin (22), betulinic acid (23), sinapyl (24), syringin (25), caffeic acid (26) and ursolic acid (27). The GI50 values of compounds 17, 18 and 20 to HepG2 cell line were 7.80, 7.08 and 4.99 µg/mL, respectively. 3. Thirteen were isolated from 95% ethanolic extract of the aerial parts of E. vestia for the first time. They were determined to be bergapten (28), xanthotoxin (29), isopimpinellin (30), esculetin (31), scopoletin (32), daphnetin (33), marmesin (34), skimmianine (35), confusameline (36), dictamine (37), obacunone (38), limonin (39) and p-hydroxy phenyl aldehyde (40). 4. The structures, biological activities, biotransformation and chemical syntheses of eudesmane-12, 6-olides from the Asteraceae during 1990-2007 were reviewed.

五种毛壳霉属和曲霉属真菌代谢产物及细胞毒活性

毛壳霉属(Chaetomium)和曲霉属(Aspergillus)真菌产生多种具有生物活性的化合物。为系统阐明两属微生物的次生代谢物，对三种毛壳霉、两种曲霉真菌分别进行固态发酵，以色谱和波谱技术研究发酵物中的成分，分离鉴定了51个化合物，其中23个为新化合物，测试了部分化合物对肿瘤细胞的活性。 1、从螺卷毛壳霉（C. cochloides）固态发酵物中分离鉴定了11个化合物，3个新化合物为螺卷毛壳霉素A~C（1~3）。化合物1、3及dethio-tetra (methylthio) chetomin（4）对Bre-04、Lu-04和N-04细胞株生长抑制的GI50值为0.05~7.0 μg/mL。 2、从印度毛壳霉（C. indicum）固态发酵物中鉴定的三个异喹啉生物碱印度毛壳霉素A~C（12~14）代表两类骨架新颖的异喹啉生物碱。 3、从巴西毛壳霉（C. brasiliense）固态发酵物中鉴定了11个化合物，其中Mollicellins I~J（15~16）、2-Hydroxymethyl-6-methylmethyleugenin（19）为新化合物。化合物16和Mollicellin H（18）对Bre-04、Lu-04、N-04细胞株生长抑制的GI50值在2.5~8.6 μg/mL。 4、从土曲霉（A. terreus）固态发酵物中鉴定了18个化合物。5个为新化合物为Terretonin A~D（24~27）和Asterrelenin（28），24~27为二倍半萜化合物，28为吲哚生物碱。 5、从杂色曲霉(A. versicolor）固态发酵物中鉴定了16个化合物。9个新的化合物Brevianamides K~N (40~43)、Averins A~C (44~46)和Glyanphenines A~B (47~48)代表三种类型的生物碱。 6、综述了1997－2007年间新的二倍半萜的研究进展。 The fungi of the genera Chaetomium and Aspergillus produce various secondary metabolites with biological activities. In order to systematically study the secondary metabolites, the solid-state fermented rice culture of three species of Chaetomium and two of Aspergillus were chemically studied. By the means of chromatograhy and spectroscopy, 55 compounds were isolated and identified, among of them 23 were new ones. The biological activities of some compounds were investigated. 1. From the fungus C. cochliodes, three new epipolythiodioxopiperazines, chaetocochins A-C (1-3) were isolated, together with 8 known ones (4-11). Compounds 1, 3 and 4 showed growth inhibitory effects against cancer cell lines Bre-04, Lu-04 and N-04 with GI50 values from 0.05 to 7.0 μg/mL. 2. Three novel isoquinolines Chaetoindicins A-C (12-14) were isolated and identified from the fungus C. indicum. Chaetoindicin A, Chaetoindicins B-C represented two classes of novel carbon skeletons. 3. Three new compounds, Mollicellins I-J (15-16), and 2-hydroxymethyl-6-methylmethyleugenin (19), were isolated from C. brasiliense. Compound 16 and Mollicellin H (18) showed growth inhibitory effects against cancer cell lines Bre-04, Lu-04 and N-04 with GI50 values from 2.5 to 8.6 μg/mL. 4. Eighteen compounds were isolated from the fungus A. terreus. Terretonin A-D（24 - 27）and Asterrelenin（28） are new compounds belonging to sesterterpoids and indole-ralated alkaloid, respectively. 5. From the fungus A. versicolor, sixteen secondary metabolites, including nine new ones, Brevianamides K-N (40-43), Averins A-C (44-46), and Glyanphenines A-B (47-48), were isolated and identified. Brevianamides K-N (40-43), Averins A-C (44-46), and Glyanphenines A-B (47-48) represented three classes of alkaloids. 6. New sesterterpenes and their bioactivities reported from 1997 to 2007 were summarized.

四种药用植物的化学成分及芍药苷的微生物转化研究

本学位论文共有5章。第一章报道白芍的化学成分及芍药苷的微生物转化研究成果；第二章报道天山雪莲的化学成分研究；第三章报道两面针的化学成分研究；第四章报道通关藤的化学成分研究成果；第五章概述了花椒属植物中最近十年报道的新化合物及药理研究情况。 在第1章的第一部分报道了白芍（Paeonia lactiflora Pall.）的化学成分。我们采用正、反相硅胶柱层析等各种分离方法，从白芍的干燥根中共分离出14个化合物，其中1个为新化合物，其结构通过波谱分析证实为没食子酰白芍苷，另外还有2个为首次从该植物中分离得到。第二部分报道了芍药苷的微生物转化生产芍药苷代谢素-I的研究，从15株厌氧菌中筛选出10株有转化活性的菌株，其中短乳杆菌Lactobacillus brevis AS1.12的转化活性最好，对其转化条件进行了初步的筛选，确定了相对合理的转化工艺。 在第2章报道了天山雪莲（Saussurea involucrate Kar.et Kir.）全草乙醇提取物化学成分的分离纯化和结构鉴定。通过正、反相硅胶柱层析等分离纯化和MS、NMR等波谱解析，共分离鉴定了28个化合物，结构类型分属于黄酮、倍半萜和木脂素等，其中2个新倍半萜化合物的结构分别表征为6α-羟基云木香酸6-β-D-吡喃葡萄糖苷和11βH-11,13-二氢去氢云木香内酯8α-O-(6′-乙酰)-β-D-吡喃葡萄糖苷。 第3章报道了两面针（Zanthoxylum nitidum （Roxb.）DC.）干燥根的乙醇提取物化学成分的分离纯化和结构鉴定。通过正、反相硅胶柱层析等分离纯化和MS、NMR等波谱解析以及X-射线单晶衍射，共分离鉴定了16个生物碱，结构类型分属于苯并啡啶类、喹啉类和阿朴啡类等，其中2个新苯并啡啶类生物碱的结构分别表征为二聚双氢两面针碱和丙酮基双氢崖定椒碱。 第4章报道了通关藤（Marsdenia tenacissima (Roxb.) Wight et Arn.）水提取物化学成分的分离纯化和结构鉴定。通过正、反相硅胶柱层析等分离纯化和MS、NMR等波谱解析以及X-射线单晶衍射，共分离鉴定了14个化合物，结构类型均属于C21多羟基甾醇，其中4个新化合物tenacigenoside A, tenacigenoside B, tenacigenoside C和tenacigenoside D的结构分别表征为3-O-6-deoxy-3-O-methyl-β-D-allopyranosyl-(1→4)-β-D-oleandropyranosyl-17β-tenacigenin B (62), 3-O-2,6- dideoxy-4-O-methyl-D-lyxo-hexopyranosly-11α-O- methylbutyryl-12β-O-acetyl-tenacigenin B (63), 3-O-6-deoxy-3-O-methyl-β-D- allopyranosyl-(1→4)-β-D-oleandropyranosyl-11α-O-tigloyl-tenacigenin C (64)和3-O-6-deoxy-3-O-methyl-β-D-allopyranosyl-(1→4)-β-D-oleandropyranosyl-11α-O-2- methylbutyryl-tenacigenin C (65)。 第5章概述了花椒属植物的化学成分及药理活性研究进展。 This dissertation consists of 5 chapters. The first chapter elaborate the phytochemical investigation of Paeonia lactiflora Pall., and microbial transformation of paeoniforin. The second, third and four chapters elaborate the phytochemical investigation of Saussurea involucrate Kar.et Kir., Zanthoxylum nitidum (Roxb.) DC. and Marsdenia tenacissima (Roxb.) Wight et Arn., respectively. Chapter 5 is a review on chemical constituents and bioactivities of Zanthoxylum species. The part one of chapter 1 focus on the isolation and identification of chemical constituents from P. lactiflora. Fourteen compounds were isolated from the roots of P. lactiflora by repeat column chromatography over normal and reversed phase silica gel. Among them, one is a new compound and the structure was suggested as galloyl-albiflorin by spectral evidence. In addition, two compounds were firstly reported in this plant. The part 2 is about microbial transformation of paeoniforin. Chapters 2, 3 and 4 were isolations and identifications of chemical constituents from S. involucrate, Z. nitidum and M. tenacissima, respectively. From the aerial parts of S. involucrate, 28 compounds including 7 flavonoids and 13 sesquiterpenoids were isolated and identified. Among them, 2 new compounds were characterized as 6α-hydroxycostic acid 6-β-D-glucoside and 11βH-11,13-dihydrodehydro- costuslactone 8α-O-(6'-acetyl)-β-D-glucoside, respectively, by means of spectroscopic analysis. Otherwise, 11 ones were firstly reported from this plant. The third chapter is about the phytochemical investigation of Z. nitidum. Sixteen compounds were isolated and identified. Among them, 2 new benzophenanthridine alkaloids were characterized as 8-acetonyldihydrofagaridine and 1,3-bis(8-dihydronitidinyl)-acetone by spectroscopic analysis. The fourth chapter is about the phytochemical investigation of M. tenacissima. Fourteen compounds were isolated and identified. Among them, 4 new compounds, tenacigenosides A~D, were characterized as 3-O-6-deoxy-3-O-methyl-β-D-allopyranosyl-(1→4)-β-D-oleandropyranosyl-17β- tenacigenin B, 3-O-2,6-dideoxy-4-O-methyl-D-lyxo-hexopyranosly-11α-O-methyl butyryl-12β-O-acetyl-tenacigenin B, 3-O-6-deoxy-3-O-methyl-β-D-allopyranosyl- (1→4)-β-D-oleandropyranosyl-11α-O-tigloyl-tenacigenin C, and 3-O-6-deoxy-3-O- methyl-β-D-allopyranosyl-(1→4)-β-D-oleandropyranosyl-11α-O-2-methylbutyryl- tenacigenin C. Chapter 5 is a review on recent progress in bioactive constituents from plants of Zanthoxylum species.

天然产物Standishinal 的首次全合成研究

具有1，1，4α-三甲基氢化芴骨架结构的天然三环二萜化合物自然界中不常见。在该类化合物中，Standishinal 具有良好的芳香化酶抑制活性和细胞毒活性。迄今未发现有Standishinal 的全合成报道，因此，我们对Standishinal 的全合成进行了探索，在该过程中得到以下实验结果： 1. 发现MSA/P2O5、MSA 在无溶剂条件下，25 °C 时烷氧基苯即可实现向苯酚的转化，但在CH3NO2 中，温度升高至80 °C 并未发生反应。 2. 烷氧基苯或对溴苯酚与α-环香叶酸在不同温度下以MSA/P2O5、MSA、PPA为催化剂以CH3NO2 为溶剂或以BF3·Et2O为催化剂时均不发生Friedel-Crafts酰化反应。 3. 对溴苯酚与香叶酸在p-TsOH 催化作用下发生了香叶酸向α-环香叶酸环化、α-环香叶酸环与对溴苯酚的酯化，得到了唯一产物α-环香叶酸对溴苯酯，产率68%。 Standishinal is one of tricyclic-diterpenes possessing the uncommon 1, 1,4a-trimethylhydrofluorene skeleton. Standishinal possesses cytotoxic and aromataseinhibitory activities. Till now, no synthesis of standishinal has been reported. Inattempt to synthesize standishinal, the following phenomenon were observed: 1. Alkyloxybenzenes could be transformed into corresponding phenol at 25 °C inthe presence of MSA/P2O5 or MSA under solvent free condition. ButAlkyloxybenzenes are stable in presence of MSA/P2O5 or MSA in CH3NO2 even at 80 °C. 2. Friedel-Crafts acylation of alkyloxybenzenes and p-bromophenol withα-cyclogeranic acid could not be realized under catalysis of MSA/P2O5, MSA or PPAin CH3NO2, or under catalysis of BF3·Et2O without CH3NO2. 3. The reaction of 4-bromaophenol and geranic acid in the presecnce of p-TsOHafforded 4-bromophenol α-cyclogeranoate in which cyclization of geranic acid toα-cyclogeranic acid was followed by esterification of α-cyclogeranic acid with p-bromophenol.

几种中药材的化学成分及其定性定量检测方法研究

本论文由四部分组成。第一部分报道了佛手参提取物的化学成分研究，建立了活性成分含量测定的高效液相测定和指纹图谱研究，采用液质联用技术鉴定了主要色谱峰；第二部分报道了丹参及其复方制剂的特征图谱研究；第三部分探讨了两面针生物碱的电喷雾质谱裂解规律，并采用液质联用技术分离鉴定了提取物中的多种生物碱。第四部分概述了液质联用在药物代谢研究中的运用。 第一部分包括第一、第二和第三章。第一章针对佛手参（Gymnadeniaconopsea）块茎的甲醇提取物，采用大孔树脂和反相硅胶柱层析等各种分离方法，共分离鉴定出4 个化合物，通过波谱分析将它们的结构确定为dactylorhin B (1)、loroglossin (2)、dactylorhin A (3)和militarine (4)。这4 个化合物均是首次从佛手参中分离得到的琥珀酸葡萄糖苷类成分。第二章采用高效液相色谱法对西藏、四川、河北、青海和尼泊尔等不同地区产的十个佛手参样品进行腺嘌呤核苷和对羟基苯甲醇的定量分析，结果表明这2 个成份可视为佛手参的特征成分，但也注意到产地不同该2 个特征成分的含量也有所不同。第三章采用标准中药指纹图谱相似度计算软件，以10 个佛手参样品HPLC 图谱的平均值为相似性评价对照模板，对10 个样品进行了相似度评价，并经液质联用分析指认了7 个共有峰，分别为腺嘌呤核苷(1)、对羟基苯甲醇(2)、对羟基苯甲醛(3) 、dactylorhin B(4) 、loroglossin(5)、dactylorhin A(6)和militarine(7)。 第二部分包括第四、第五、第六和第七章。第四章运用电喷雾质谱检测了对照药材和五个不同产地的丹参药材中脂溶性和水溶性成分，系统地探讨了多种成分的电喷雾质谱规律，并以对照药材为标准建立了特征指纹图谱。五个产地的药II材通过与对照药材相对比，采用聚类分析的方法，得到了定性的鉴别与判断。并采用液质联用技术对丹参药材提取液中的化学成份进行分析，推测了九个特征峰，并对六样品的液相色谱图进行了聚类分析。第五章探讨了三七皂苷的电喷雾质谱电离和裂解规律，并采用电喷雾质谱法对三七标准药材，血通片中的皂苷成分进行了分析。第六章运用电喷雾质谱研究复方丹参片提取液的特征图谱，并和单味药材丹参和三七的特征图谱进行了对比研究。并运用HPLC-ESI MSn 分析鉴定了复方丹参片提取液中的化学成分，推测了12 个色谱峰。第七章总结了电喷雾质谱和液质联用技术在丹参药材，三七药材及复方丹参制剂中的运用的优势和局限性。 第三部分（第八章）研究了两面针生物碱中二氢白屈菜红碱(1)、二氢两面针碱(2)、8-酮基二氢白屈菜红碱(3)、8-丙酮基二氢两面针碱(4)、两面针碱(5)、和1,3-二(8-二氢两面针碱)丙酮(6)等六个苯并菲啶型生物碱的电喷雾质谱裂解规律，其中二氢两面针碱和二氢白屈菜红碱，8-丙酮基二氢两面针碱和8-酮基二氢白屈菜红碱是两对二个甲氧基分别在C-9 和C-10，C-10 和C-11 的同分异构体。实验结果表明，在相同的碰撞能下，这类位置异构体的ESI MS2 质谱二级碎片离子的相对峰度存在很大差异，这可以用于区分该类同分异构体，采用液-质联用可以对两面针的总生物碱提取物中的这些同分异构体加于区分。同时在本实验采用的液相色谱条件下，多种生物碱得到较好的分离，通过和对照品的保留时间，紫外吸收光谱及电喷雾质谱图对照，鉴定了11 个主要色谱峰。 第四部分（第九章）对液质联用技术在药物代谢中的运用进行了综述。 This dissertation consisted of four sections. The first two sections elaborated thephytochemical investigation of the rhizomes Gymnadenia conopsea R. Br., methoddevelopment for rapid identifying and qutifying the chemical condtituent of thistibetant medicine, and the chemical fingerprint analysis of rhizomes of G. conopsea,Salviae miltiorrhiza and P. notoginseng. The third section studied the fragmentationmechanism of six alkaloids from Zanthoxylum nitidium and method development forrapid identifying varieties of alkaloids from the extract of this herbal medicine. Thefourth section reviewed HPLC- MS method in drug metabolism studies. The first section consisted of chapters 1, 2, 3. Chapter 1 elaborated the phytochemicalinvestigation of Gymnadenia conopsea R. Br. Four succinate derivative esters wereisolated from the methanol extract of the rhizomes of G. conopsea through repeatedcolumn chromatography on normal and reversed phase silica gel, their structures weredetermined by ESI-MS, 1D and 2D NMR evidence. They were firstly discoveredfrom this species. In chapter 2, a high-performance liquid chromatography.diodearray detection (HPLC-DAD) method has been firstly developed for quantitation oftwo characteristic constituents, adenosine and 4-hydroxybenzyl alcohol, from theextract of rhizomes of G. conopsea. All 10 samples of G. conopsea contained differentamount of adenosine and 4-hydroxybenzyl alcohol. Adenosine and the4-hydroxybenzyl alcohol can be applied in identification and quality control for theroots of G. conopsea. In chapter 3, a high-performance liquid chromatography.diodearray detection.tandem mass spectrometry (HPLC-DAD-MSn) method has been firstly developed for chemical fingerprint analysis of rhizomes of G. conopsea andrapid identification of major compounds in the fingerprints. Comparing the UV andMS spectra with those of authentic compounds, seven main peaks in the fingerprintswere identified as adenosine, 4-hydroxybenzyl alcohol, 4-hydroxybenzyl aldehyde,dactylorhin B, loroglossin, dactylorhin A and militarine. The Computer AidedSimilarity Evaluation System for Chromatographic Fingerprint of TraditionalChinese Medicine (CASES) was employed to evaluate the similarities of 10 samplesof the rhizomes of G. conopsea collected from Sichuan, Qinghai and Hebei provincesand Tibet autonomous region of China, and Nepal. These samples from differentsources had similar chemical fingerprints to each other. The second section consisted of chapters 4, 5, 6 and 7. In chapter 4，both thecharacteristic spectra of liposoluble tanshinones and aqueous-soluble salvianolic acidswere established by the electrospray ionization mass spectrometry (ESI MS)technique and the differences between standard and crude rhizomes of Salviaemiltiorrhiza Bge. from 5 sources were analyzed. The law of electrospray ion trap mass(ESI ITMS) of typical tanshinones and salvianolic acids is studied.The analysis of the chemical constituent of rhizomes of Salviae miltiorrhiza Bge. byliquid chromatography coupled with mass spectrum (LC/MS) technique wasestablished，and the distances among standard herb and crude herb from 5 sourceswere calculated by clustering analysis. According the DAD spectra and MS2 data，9tanshinones could be speculated. In chapter 5, the character spectra of total saponinsin P. notoginseng extracts were established by ESI ITMS and selective ion monitoring(SIM) technology. The law of notoginsenosides by ESI MS2 was studied. In chapter 6,the characteristic spectra of Compound Danshen Tablet established and compared byESI-MS and HPLC/DAD/MS, 6 known tanshinones and 3 saponins were speculated.In chapter 7, the advantage and disadvantage of the strategy, using the ESI ITMS andLC/MS techniques for study of characteristic spetra of danshen and Compound Danshen Tablet, were summerized. The third section (chapter 8) studied the fragmentation mechanism of six alkaloids,dihydronitidine, dihydrochelerythrine, 8-acetonyl dihydronitidine,8-acetonyldrochelerythrine, nitidine and 1,3-bis (8-dihydronitidinyl)-acetone, by ESIMSn. Tandem mass spectrometry experiments indicated that different substitutionsites of the methoxyl groups at C-9 and C-10 or at C-10 and C-11 determined thedifferent abundances of the MS2 fragmentation ions using the same collision energy.According to the different abundances of MS2 product ions, positional isomericbenzo[c] phenanthridine alkaloids can be differentiated. Moreover, ten constituents inthe crude alkaloids extract from the roots of Zanthoxylum nitidium were rapidlyidentified by high-performance liquid chromatography coupled with tandem massspectrometry (HPLC-MSn), through comparing the retention times and ESI MSn spectra with the authentic standards. The fourth section (chapter 9) is a review on HPLC-MS method development in drug metabolism studies.

五枫藤、小驳骨和川西茶藨的化学成分研究

八月瓜属植物五枫藤(Holboellia latifolia Wall.)和驳骨草属植物小驳骨(Gendarussa vulgaris Nees)均为药用植物, 前者化学成分研究不深入, 后者的化学成分未见报道。川西茶藨(Ribes takare D. Don)为茶藨子属植物, 没有化学成分的报道。本论文对三个植物的化学成分和活性成分进行了研究, 主要通过色谱方法分离得到了48 个化合物, 采用波谱分析或与已知标准品对照等手段鉴定了它们的结构, 其中有1 个新的原小檗碱类化合物和3 个新的联苯类化合物,发现了具有细胞毒活性和α-葡萄糖苷酶抑制活性的化合物。1、从五枫藤地上部分的95%乙醇提取物中分离得到了12 个化合物: 五加苷K (1)、hederagenin 3-O- α-L-rhamnopyranosyl-(1→2)- α-L-arabinopyranoside (2)、β-萘乙酸(3) 、3-O-α-L-rhamnopyranosyl-(1→2)-[β-D-glucopyranosyl-(1→3)]-α-L-arabinopyranosyl oleanolic acid 28-O-α-L-rhamnopyranosyl-(1→4)-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl ester (4) 、3-O- α-L-rhamnopyranosyl-(1→2)-O- β- D-glucopyranosyl-(1→2)- α-L-arabinopyranosyl oleanolic acid (5) 、3-O-( β-D-glucopyranosiduronic acid)-oleanolic acid 28-O- β-D-glucopyranoside (6)、lup-20(29)-en-3-one (7)、lupeol (8)、β-谷甾醇(9)、齐墩果酸(10)、乌苏酸(11)、β-胡萝卜苷(12)。化合物1 对Lu-06、N-04 和Bre-04 癌细胞株的GI50 分别是0.77µg/mL、1.26 µg/mL 和1.55 µg/mL, 化合物2 对N-04 癌细胞株的GI50 为2.44 µg/mL。2、从小驳骨地上部分的95%乙醇提取物中分离得到了1 个原小檗碱类新化合物13-hydroxyl gusanlung A (25), β-谷甾醇(9)、齐敦果酸(10)、β-胡萝卜苷(12)、棕榈酸(1-)甘油酯(13)、棕榈酸(14)、阿苯哒唑(15)、阿苯哒唑砜(16)、阿苯哒唑亚砜(17)、aurantiamide acetate (18)、华良姜素(19)、芫花素(20)、(-)-丁香树酯醇(21)、gusanlung B (22) 、eupteleasaponinsⅤ acetate (23)、gusanlungA (24)、刺五加苷E (26)、岩白菜素(27)、咖啡酸(28)。化合物25 对肝癌细胞株(HepG2) 的GI50 为2.08 µg/mL。3、从川西茶藨地上部分的95%乙醇提取物中分离鉴定了22 个化合物: β-谷甾醇(9) 、β- 胡萝卜苷(12) 、O-acetyloleanolic aldehyde (29),4,7,8-trimethoxy-2,3-methylenedioxydibenzofuran (30) 、3', 5-dimethoxy-3, 4-methylenedioxybiphenyl (31) 、桦木醇(32) 、6,7-dimethoxy-1-methyl-3,4-dihydroquinolin-2-one (33)、3'-hydroxy-5-methoxy-3,4-methylenedioxybiphenyl (34) 、7-hydroxy-4,8-dimethoxy-2,3-methylenedioxydibenzofuran (35)、桦木醛(36)、没食子酸(37) 、6β- 羟基-4- 烯-3- 酮- 豆甾醇(38) 、5α, 8α-epidioxy-(22E,24R)-ergosta-6, 22-dien-3β-ol (39)、verrucofortine (40)、6-methoxycalpogoniumisoflavone A (41)、2-羟基二苯甲酮(42)、桦木酸(43), 3, 5-二甲氧基苯甲酸-4-O-β-D-吡喃葡萄糖苷(44)、洋芹素(45)、刺槐素(46)、水杨酸(47)、洋芹素-5-O- β-D-葡萄糖苷(48), 化合物30、31 和35 为新的联苯化合物。化合物30的α-葡萄糖苷酶抑制率为10.2% (1.00 mg/mL); 化合物35 的抑制率为17.2% (1.00mg/mL)。4、综述了1960 年以来原小檗碱类化合物药理活性研究进展。 Plants Holboellia latifolia Wall and Gendarussa vulgaris Nees, are used as folkmedicine. Ribes takare D. Don belongs to the genus Ribes. The three plants have notbeen chemically studied in detail. Chemical and bioactive study of three plants led tothe isolation of 48 compounds by chromatography. Their structures were elucidatedon the basis of spectroscopic evidence or comparison with authentic samples. Amongthe 48 componds isolated one protoberberine alkaloid and three biphenyls are newones. Cytotoxic and α-glucosidase inhibitory compounds had been found.1. Twelve compounds were isolated from the 95% ethanol extract of the aerial partof H. latifolia Wall. They were characterized as fellow: eleutheroside K (1),hederagenin-3-O- α-L-rhamnopyranosyl-(1→2)- α-L-arabinopyranoside (2),2-naphthyl acetic acid (3),3-O-α-L-rhamnopyranosyl-(1→2)-[β-D-glucopyranosyl-(1→3)]-α-L-arabinopyranosyl oleanolic acid 28-O-α-L-rhamnopyranosyl-(1→4)-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl ester (4), 3-O- α-L-rhamnopyranosyl-(1→2)-O- β- D-glucopyranosyl-(1→2)- α-L-arabinopyranosyl oleanolic acid (5),3-O-( β-D-glucopyranosiduronic acid)-oleanolic acid-28-O- β-D-glucopyranoside (6),lup-20(29)-en-3-one (7), lupeol (8), β-sitosterol (9), oleanolic acid (10), ursolicacid (11), and β-daucosterol (12). Compound 1 showed moderate cytotoxicity againstLu-06 (GI50, 0.77 µg/mL), N-04 (GI50, 1.26 µg/mL) and Bre0-4 (GI50=1.55 µg/mL)and compound 2 showed moderate cytotoxicity against N-04 (GI50=2.44 µg/mL).2. A new protoberberine alkaloid, 13-hydroxyl gusanlung A (25), was isolated fromthe aerial part of Gendarussa vulgaris Nees, together with β-sitosterol (9), oleanolicacid (10), β-daucosterol (12), glycerol monopalmitate (13), palmific acid (14),albendazole (15), albendazole sulphone (16), albendazole sufloxide (17), aurantiamideacetate (18), kumatakenin (19), genkwanin (20), (-)-syringaresinol (21), gusanlung B(22), eupteleasaponinsⅤ acetate (23), gusanlung A (24), eleutheroside E (26),bergenin (27) and caffeic acid (28). Compound 25 showed cytotoxicity against HepG2 cells (GI50, 2.08 µg/mL).3. Phytochemical study of the Ribes takare D. Don led to the isolation of three newbiphenyls, 4,7,8-trimethoxy-2,3-methylenedioxydibenzofuran (30), 3', 5-dimethoxy-3,4-methylenedioxybiphenyl (31) and 7-hydroxy-4,8-dimethoxy-2,3-methylenedioxydibenzofuran (35), along with nineteenknown compounds, β-sitosterol (9), β-daucosterol (12), O-acetyloleanolic aldehyde(29), betulin (32), 6,7-dimethoxy-1-methyl-3,4-dihydroquinolin-2-one (33),3'-hydroxy-5-methoxy-3, 4-methylenedioxybiphenyl (34), betulinic aldehyde (36),gallic acid (37), stigmast-4-en-6β-ol-3-one (38), 5α, 8α-epidioxy-(22E, 24R)-ergosta-6,22-dien-3β-ol (39), verrucofortine (40), 6-methoxycalpogonium isoflavone A (41),2-hydroxybenzophenone (42), betulinic acid (43), 3,5-dimethoxygallic acid-4-O- β-D-glucopryranoside (44), apigenin (45), acacetin (46), salicylic acid (47) andapigenin-5-O- β-D-glucopryranoside (48). α-Glucosidase inhibitory rates ofcompound 30 and 35 were respectively 10.2% and 17.2% at a concentration of 1.00 mg/mL).4. Pharmacological activities of protoberberines were summarized.

二萜生物碱植物的化学成分研究

本论文由三个部分组成。第一部分是综述性文章-天然C19-二萜生物碱的结构及其核磁共振波谱。这篇综述参考160余篇文献，将迄今为止发表的230余个天然C19-二萜生物碱按其结构和波谱特征，分为10个类型。重点叙述了每个类型的结构和核磁共振波谱，对一些立体构型与13C-NMR化学位移的关系进行了讨论。最后以表格的形式报导了每个天然C19-二萜生物碱的结构及其植物来源，并把其中170余个收集到的13C-NMR化学位移数据整理成表，作为手册，对鉴定此类化合物大有帮助。第二部分是实验论文--弯啄乌头的化学成分研究。重点报导了弯啄乌头(Aconitum campylorrhynchun Hand-Nazz.)中分离到的一个新生物碱--8-乙酰都啦碱(8-acetyldolaconine)的结构鉴定，并根据DEPT、COSY、COLO等对1H-NMR和13C-NMR的所有信号归属都进行了指定。通过各种图谱还发现这个生物碱的A环同时存在着椅式和船式两种构象导购体。另外还报导了从同一植物的生物碱部分分离到的两个二萜生物碱，它们都是嘟啦碱和aconosine.在非生物碱部分还分离到三个化合物，经鉴定证明是β-谷甾醇、棕榈酸和香豆酸。第三部分也是实验论文--展毛翠雀花中生物碱成分的研究。报导了从展毛翠雀花(Delphinium Kamaonense Var.glabrescens W. T. Wang)中初步分离到的两个生物碱。经IR、NS、1H-NMR和13C-NMR等方法确定结构，证明其中一个是新化合物，命名为展毛翠雀碱(glabredelphinine)。另外一个是前几年发现的二萜生物碱tatsiensine。这两个化合物的A环上都有碳-碳不饱和键，这在天然C19-二萜生物碱中还比较少见。This master's thesis consists of three parts. The first part is a review: The relationship between the chemical structures of natural C19-diterpene alkaloids and their NMR chemical shift data. The C19-diterpene alkaloids were divided into ten types according to their structures and NMR data. Some relationaships between configuration and NMR chemical shift have been discussed. The structures and distributions of more than 230 known C19-diterpene alkaloids and the assignments of 13C-NMR data of the 170 alkaloids of them were listed in tables. The second part is an experimental paper, Studies on the chemical constituents of Aconitum campylorrhynchum. Six compounds were isolated from the roots of title plant. One of them is a new alkaloid. It has been elucidaled as 8-acetyldolaconine by means of IR, MS, 1H-NMR,13C-NMR,DEPT,COSY,COLOC and on the basis of identification with aconosine by saponification. Other five compounds were idenfified as dolaconine, aconosine,β-sitosterol, palmitic acid and p-coumaric acid. The third part is also experimental paper: The alkaloids of Delphinium Kamaonense var.glabrescens. Two alkaloids were isolated from the roots of title plant. One of them, C22H22O6N,m.p.201-3℃,named glabredelphinine, was proved to be a new compound. Its structure has been established as 2(3)-dehydro-6-demethyl-18-delcosine by means of IR, MS, 1H-NMR and 13C-NMR.The other one was identified as tatsiensine by comparising of IR, MS, 1H-NMR and 13C-NMR data with that from the literatute.

千金子和暖地大叶藓的化学成分研究

本论文由三部分共4 章组成。第一部分阐述了大戟科大戟属传统中药千金子（Euphorbia lathyris L.）化学成分、生物学活性以及千金子化学成分的HPLC、UPLC-MS、GC-MS 分析成果。第二部分介绍了民族药材暖地大叶藓（Rhodobryum giganteum (Schwaegr.) Par.）的化学成分研究和结构鉴定。第三部分概述了大戟属 植物中大环二萜酯的研究进展。 第一章包括1－3 节。在第1, 2 节中报道了千金子（Euphorbia lathyris L.）95% 乙醇提取物的化学成分分离鉴定。我们采用正、反相硅胶柱层析、重结晶等各种分离方法，凭借MS、IR、NMR、X-ray 等现代仪器手段,从中共分离鉴定22 个化合物。其中8 个是高活性化合物前体-续随子烷型大环二萜及3 个巨大戟烷型二萜，还有香豆素、生物碱、甾体等类型，其中完成对5 个大环二萜酯构型的确认，对2 个二萜酯构型进行了修正。第3 节中介绍对千金子化学成分的细胞毒性、α-葡萄糖苷酶抑制活性、P-gp 表达抑制活性的模型筛选结果。 第二章包括3 节，第1 节报道不同产地千金子高效液相色谱定量分析结果。第2 节介绍了各大环二萜酯的HPLC-MS/MS 的分析结果，并且对其质谱裂解规律、UPLC-MS 快速鉴定方法做了进一步讨论。第3 节介绍了千金子挥发油成分分析。采用传统水蒸气蒸馏方法提取千金子中的挥发油，并经气相色谱－质谱联用（GC-MS）技术共分离鉴定出 49 个化合物，占挥发油总量的90.48％。 第三章包括1, 2 两节，第1 节报道了暖地大叶藓化学成分。采用正、反相硅胶，凝胶柱层析等各种分离方法和MS、IR、NMR 等解析手段，共分离鉴定10个化合物，其中一个环肽化合物为新化合物。第2 节介绍了暖地大叶藓挥发油成分分析，共分离鉴定出 52 个化合物，占其挥发油总量的85.67％。 第四章概述了大戟科大戟属植物中大环二萜酯的研究进展。 This dissertation consists of three parts. In the first part, it is elaborated that the phytochemical investigation from the traditional Chinese medicine: seeds of Euphorbia lathyris L.. Biological activity and constituents analysis by HPLC、UPLC-MS、GC-MS were reported. In the second part, it is discussed that the chemical constituents were isolated and identificated from minority nationalitical herb－Rhodobryum giganteum (Schwaegr.) Par.. The third part is a review about the progress of studies on macrocyclic diterpenes from Euphorbia. The first part is composed of 1－3 sections. The section 1and 2 is focused on the isolation and identification of chemical constituents from seeds of E. lathyris. 22 compounds were isolated from the seeds of E. lathyris. by isolation methods of column chromatography (silica gel, including reversed phase) and recrystallisation on the basis of spectroscopic methods including IR, MS, NMR and X-ray. In 8 macrocyclic and 3 ingenane diterpenes, the relative configuration of 5 macrocyclic diterpenes were confirmed, in which 2 were amended. In the third section, cell cytotoxic activity, restraining activity of α-Glucosidase and multidrug resistance (MDR) reversing activity about P-gp were tested. 5 potential revsering reagents were found. The second part is composed of 1－3 sections. In first section it is described that the quality of the chemical constituents of E. lathyris from 5 sources , which were analyzed by high-performance liquid chromatography. In addition, the fractionation rules of some macrocyclic diterpenes were discussed and Ultra Performance Liquid Chromatography/ electrospray ionization mass spectrometry (UPLC-MS) was applied for quick determination of compounds in the second section. In the third section, chemical analysis of the essential oil from seeds of E. lathyris by GC-MS were reported. The essential oil from the seeds of E. lathyris L. in Sichuan was extracted by steam distillation and 49 compounds were isolated and identified from the essential oil by gas chromatography-mass spectrometer (GC-MS). These compounds are accounted for 90.46% of the total essential oil. The second part, including section 4 and 5, is about the phytochemical investigation of R. giganteum. In the former section, ten compounds were isolated and identified. Among them, a new peptide was characterized by spectroscopic analysis including IR, MS and NMR. In the other section, 52 compounds were isolated and identified from the essential oil by gas chromatography-mass spectrometer (GC-MS). These compounds are accounted for 85.67% of the total essential oil. The third part is a review about the progress of studies on macrocyclic diterpenes from Euphorbia.

低极性化合物的电喷雾质谱检测方法及几种中藏药材的活性成分研究

本学位论文首先报道了为解决低极性化合物的电喷雾质谱（ESI-MS）分析难题而建立的一种衍生化分析方法；然后从色谱-质谱联用分析、分离纯化和结构鉴定等方面分别报道了几种中藏药材的活性成分研究。论文由下述六章组成： 第一章报道了盐酸羟胺衍生化方法在电喷雾质谱 (ESI-MS) 分析中的应用。该方法利用盐酸羟胺和羰基成肟的快速反应，建立了针对三萜酮等含酮或醛羰基低极性化合物的ESI-MS 信号增强技术。此方法不仅可应用于增强羰基化合物的ESI-MS 质谱信号，还可检测化合物中羰基的个数以及辨别涉及羰基官能团的同分异构体。此外，通过简单的氧化反应，还可将该方法拓展到三萜醇、甾醇等含羟基的低极性化合物，增强它们的ESI-MS 信号。对比已报道的相关ESI-MS 增强质谱信号的衍生化方法，此方法有经济、实用、快速和简便的显著特点。 第二章是关于野生羌活及其栽培品种化学成分的色谱-质谱联用分析。对不同产地野生羌活生长过程中活性成分的动态变化、野生羌活不同形态部位和人工栽培羌活中的活性成分含量进行了HPLC 定量分析。结果表明主要活性成分羌活醇和异欧前胡素都随生长期存在规律性变化，羌活不同形态部位中的活性成分含量也有明显不同。这些实验结果有些较好地印证了传统中医的用药理论，有些也对羌活的传统使用方法提出了新的建议。 第三章介绍了几种传统中藏药材的色谱-质谱联用及串联质谱分析。通过GC-MS 方法，从藏药材长花党参挥发油中共分离鉴定出45 个化合物；利用HPLC方法测定了该藏药材中的主要化学成分——木犀草素的含量（0.7%）；利用串联质谱技术，对西番莲和射干中的主要成分进行了快速鉴定，从西番莲中鉴定了4个黄酮碳苷；从不同产地的射干和川射干中鉴定了8 个主要异黄酮成分，其中包括一个未见报道的化合物。 第四章的内容为藏药材石莲叶点地梅的活性成分研究。从植物石莲叶点地梅(Androsace integra (Maxim.) Hand.-Mazz.) 乙醇提取物的正丁醇萃取部分共分离和鉴定了6 个化合物，利用MS 和NMR 等现代波谱学技术阐明了它们的结构：其中包括4 个三萜类化合物：分别是androsacin (1)、 ardisiacrispin A (2) 、saxifragifolin A (3) 和20(29)-lupen-3-one (4)；一个神经酰胺：4-羟基-Δ8,9(Z)-鞘氨醇-2'-羟基正二十四碳酸酰胺(5)；一个甾体类化合物：胡萝卜苷(6)。化合物1为新的13,28-epoxy-oleanane 型三萜皂苷，在其结构表征的过程中，采用LC-MS 进行糖分析，获得了值得推广的好结果。通过活性筛选发现化合物1~3 对HepG2肝癌细胞表现出不同程度的抑制活性，其中化合物2 活性最好，其IG50 为1.65μg/mL。 第五章是关于一些传统中藏药材的农药活性筛选。利用Syngenta 公司的活性筛选平台对68 种传统中藏药材醇提物进行了抗菌和除草的生物源农药活性筛选。结果表明所筛选的68 种植物提取物中，共有14 种样品表现出明显的除草/杀虫活性，其中水母雪莲花、松萝和茯神木等植物提取物还具有多种生物活性。活性成分还有待进一步追踪分离、纯化和结构鉴定。 第六章为文献综述，概述了羌活药材的研究进展。对羌活属及药用羌活植物从分类学、本草学、品质评价、人工栽培、化学成分及药理作用等方面进行了文献归纳和总结。 In this dissertation, an electrospray ionization mass spectrometry (ESI-MS) signal enhancement method, as well as the work of bioactive components study, HPLC-MS/MS application, bioassay screening, chromatograph separation and structure identification of the metabolites in several medicinal herbs have been reported. First chapter expounded a rapid, simple ESI-MS sensitivity enhancement method for detecting carbonyl groups in natural products has been developed by using hydroxylamine hydrochloride (NH2OH·HCl) as a derivatization reagent. We use the oxime formed during the derivatization reactions and its Beckmann rearrangement intermediates as a means of detecting the carbonyl groups originally present in these triterpenoids. In comparison with other derivatization methods in the literature, this method is simple, specific and can be used to detect carbonyl groups in triterpenoids which have low polarity and are poorly or non-ionizable. Moreover, it can also be used to detect hydroxyl groups by using the Dess-Martin periodinane (DMP) to convert primary and secondary hydroxyls into carbonyl groups. Chapter 2 reported an HPLC-MS method for analyzing the main bioactive compounds in both wild and cultured Notopterygium incisum. The results indicated that the main bioactive compounds varied through different seasons regularly, and in different commercial parts of this herb the content of these compounds also differed from each other. The quantitative analysis results showed that in the traditional commercial parts, the content of main chemical constitutes in Silkworm Notopterygium, Bamboo Notopterygium and Irregular-nodal Notopterygium are higher than that in Striped Notopterygium. This result is tally with the traditionally concept that the quality of Notopterygium, Bamboo Notopterygium and Irregular-nodal Notopterygium are better than that of Striped Notopterygium, which means that the quality of rhizomes is better than main roots. The chemical constituents of cultured N. incisum is reported for the first time in this dissertation and the analysis results showed some growth curves of chemical constituents in this plant, but still left some questions unanswered. Chapter 3 discussed the GC/LC-MS analysis of the traditional Chinese medicines Codonopsis thalictrifolis, Passiflora incarnate, Belamcanda chinensis and Passiflora incarnate. The main constituent, luteolin was isolated and identified from the traditional Tibet medicine of C. thalictrifolis. The quantitative analysis by HPLC has revealed that the content of luteolin in this herb is 0.7%. GC-MS was employed to analyzed chemical constituents of the essential oil from the flower of C. thalictrifolis. More than 60 peaks were detected and 45 of them were identified by comparing their spectra with that of the standards in the database and literatures. ESI-MS/MS was used to analyze the n-butanol extract of Passiflora incarnate. Based on the information of pseudo molecular ions and fragment ions of the glycosides, four major flavone-C-glycosides have been detected and identified as 7-methoxyluteolin-6-C-β-D-glucopyranoside, vitexin, swertisin and orientin. The isoflavone compounds in theextracts of three samples of B. chinensis collected in Gansu, Sichuan and Hunan, and the extract of Iris tectorum collected in Sichuan were analyzed by using TOF-HRMS and IT-MS. From the extracts of these herbs, a new isoflavone, identified as 5’,5,6,7-tetrahydroxy-3’4’-dimethoxyl isoflavon, and 7 known ones have been identified by analyzing the fragmentation patterns and their molecular formulas given by HRMS and the tandem mass spectrometry acquired by IT-MS. Chapter 4 elucidated the isolation and identification of a new triterpene saponin, androsacin (1), along with five known compounds (2-6) were isolated from the whole plants of Androsace integra (Maxim.) Hand.-Mazz., an herb used in traditional Chinese and Tibetan medicine. The chemical structure of the new compound was established as 3β-O-{β-D-glucopyranosyl-(1→4)-O-β-D-xylopyranosyl-(1→2)-O-β-D-glucopyranosyl-(1→4)-[O-β-D-glucopyranosyl-(1→2)]-α-L-arabinopyranosyl}-16α-hydroxy-13β,28-epoxy-olean-30-al by analyzing its MS, 1D- and 2D-NMR spectra. Compound 2 was cytotoxic toward HepG2 cancer cell with the GI50 value of 1.65 μg/mL. Chapter 5 described the biogenic pesticide activity screening of 68 traditional Chinese and Tibetan medicine extractions. The intention of this study is to explore bioactive natural compounds from these traditional medicinal herbs for biogenic insecticides use. Based on Syngenta’s bioassay, 14 extractions of these traditional medicines showed pesticide activities, and some of them had multi-activities on antibacterial and insecticidal. Chapter 6 is a review on the chemical and bioactivity research progress of Notopterygium incisum and N. forbesii.

三种藏药材的化学成分研究

本学位论文报道了西藏产三种藏族传统植物药材的化学成分研究。论文由四章组成，前三章是实验部分，分别报道了尼泊尔黄堇(Corydalis hendersonii Hemsl.)、藏波罗花(Incarvillea younghusbandii Sprague)和全缘叶绿绒蒿(Meconopsis interifolia Franch.)的化学成分研究结果。从这三种青藏高原药用植物中共分离鉴定出33 个化合物，其中1 个是新化合物。第四章概述了罂粟科紫堇属植物的化学和药理研究进展。 第一章为尼泊尔黄堇的化学成分研究。通过正、反相硅胶柱色谱等分离方法从药用植物尼泊尔黄堇的地上部分共分离纯化得到12 个化合物。运用MS、1H-NMR、13C-NMR、DEPT、HMBC、NOESY 等现代波谱学方法将它们的结构鉴定为：刺罂粟碱(1) ， 普托品(2) ， 新那亭(3) ， 斯可任(4) ， tetrahydrothalifendine (5) ，9-methyl-decumbenine C (6)，tetrahydroberberrubine (7)，隐品碱(8)，α-别隐品碱(9)，6,7-methylenedioxy-1(2H)-oxoisoquinolinone (10)，6-丙酮基-5,6 -二氢血根碱(11)和β-谷甾醇(12)。其中化合物6 为新化合物，为首次发现的分子骨架上C-9 位连有甲基的苯肽异喹啉类型生物碱。另外，除化合物1 和2 外，其它9 个生物碱（3～11）均为首次从该种植物中分离得到。同时，我们还对对尼泊尔黄堇中的总生物碱进行了串联质谱分析。 第二章为藏波罗花的化学成分研究。从该药用植物的地上部分共分离得到16个化合物，通过理化常数和波谱数据鉴定为：异佛手柑内酯(1)，6-甲氧基当归素(2)，欧前胡素(3)，花椒毒内酯(4)，珊瑚菜素(5)，水合氧化前胡素(6)，rivulobirin A (7)，齐墩果酸甲酯(8)，咖啡酸甲酯(9)，银桦酸(10)，(D)-boschniakinic acid (11)，对羟基苯甲酸(12) ， tert-O-β-D-glucopyranosyl-(R)-heraclenol (13) ， 5-methoxy-8-O-β-D-glucopyranosyloxypsoralen (14)，前胡苷V(15)和苯乙醇-O-β-D-吡喃葡萄糖-(1→2)-O-β-D-吡喃葡萄糖苷(16)。所有以上化合物均为首次从该种植物中分离得到。另外我们还首次对藏波罗花挥发油的化学成分进行了气相色谱-质谱(GC-MS)联用分析，共鉴定出39 个挥发性成分。 第三章为全缘叶绿绒蒿化学成分的分离鉴定。从传统藏药材全缘叶绿绒蒿地上部分共分离纯化出8 个化合物。通过理化常数和波谱数据将他们的结构分别鉴定为：去甲血根碱(1)，β-谷甾醇(2)，3-羟基-齐墩果烷-12(13)-烯-30-酸(3)，6-丙酮基-5,6-二氢血根碱(4)，木犀草素(5)，胡萝卜苷(6)，quercetin 3-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranoside (7)和普托品(8)。其中化合物1，4 和7 为首次从该种药用植物中分离得到。 第四章为综述，总结和归纳了近年来罂粟科紫堇属植物的化学和药理研究进展。 This dissertation consists of four parts. The first, second and third parts report the studies on the chemical constituents from the medicinal plants of Corydalis hendersonii, Incarvillea younghusbandii and Meconopsis interifolia. The forth part reviews the progress of the studies on Corydalis species. The first chapter is about the isolations and identifications of alkalids from the aerial parts of C. hendersonii which is a traditional Tibetan medicine to treat febrifuge, high blood pressure and hepatitis. A new isoquinoline alkaloid, 9-methyl-decumbenine C (6), together with ten known alkaloids, stylopine (1), protopine (2), canadine (3), scoulerine (4), tetrahydrothalifendine (5), tetrahydroberberrubine (7), cryptopine (8), α-allocryptopine (9), 6,7-methylenedioxy-1(2H)-isoquinolinone (10) and 6-acetonyl-5,6-dihydrosanguinarine (11), and β-sitosterol (12) were isolated. Their structures were elucidated by spectroscopic methods. Furthermore, the total alkaloids were analyzed by ESI-MSn. The second chapter is about the isolations and identifications of chemical constituents from the aerial parts of I. younghusbandii. Sixteen compounds were isolated and purified by normal and reversed phase silica gel column chromatography. By spectral analysis, there structures were identified as isobergapten (1), sphondin (2), imperatorin (3), xanthotoxin (4), phellopterin (5), heraclenol (6), rivulobirin A (7), methyl oleanolate (8), methyl caffeate (9), grevillic acid (10), (D)-boschniakinic acid (11), 4-hydroxybenzoic acid (12), tert-O-β-D-glucopyranosyl-(R)-heraclenol (13), 5-methoxy-8-O-β-D-glucopyranosyloxypsoralen (14), decuroside Ⅴ(15), and phenylethyl-O-β-Dglucopyranosyl-(1→2)-β-D-glucopyranoside (16). All of these compounds were isolated from this plant for the first time．By the way, the chemical components of the essential oil from I. younghusbandii were analyzed by GC-MS for the first time. The third chapter is about the the isolations and identifications of the chemical constituents of M. interifolia. Eight compounds were isolated and identified as norsanguinarine (1), β-sitosterol (2), 3-hydroxyolean-12(13)-en-30-oic acid (3), 6-acetonyl-5,6-dihydrosanguinarine (4), luteolin (5), daucosterol (6), quercetin 3-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranoside (7) and protopine (8). The compounds 1, 4 and 7 were isolated from this herb for the first time. The last chapter is a review of the research progress of the studies on Corydalis species, which includes the chemical constituents in this genus and their pharmacology.

厚朴中α-葡萄糖苷酶抑制活性成分研究

本论文由三章组成。 第一章是关于厚朴中具有α-葡萄糖苷酶抑制活性成分的研究。凹叶厚朴的乙醇提取物显示了较强的α-葡萄糖苷酶抑制活性。为了确定其活性成分，在活性测试的指导下，通过溶剂萃取、树脂吸附和反复硅胶柱层析等分离方法从凹叶厚朴乙醇提取物中分离得到6 个生物碱，并用质谱和核磁共振等波谱方法分别鉴定为：木兰箭毒碱，木兰花碱，鹅掌楸碱，蕃荔枝碱，罗默碱和Lysicamine。应用小肠α-葡萄糖苷酶模型测定了它们对α-葡萄糖苷酶的抑制作用。其中，番荔枝碱和木兰箭毒碱对α-葡萄糖苷酶相对抑制活性最好，分别为60%和62%；其它四个生物碱成分对α-葡萄糖苷酶的抑制活性几乎相当，鹅掌楸碱为46%，罗默碱为51%，Lysicamine 为49%，木兰花碱为51%。 第二章报道了厚朴酚的衍生物及其对α-葡萄糖苷酶的抑制活性。根据糖苷酶抑制剂的结构特点，设计合成了一系列厚朴酚的衍生物。厚朴酚经过Mannich 反应和环氧化及开环反应制备了一系列衍生物，经活性测试发现衍生物活性与取代基关系较大，其中5,5′-diallyl-3-((bis(2-hydroxyethyl)amino)methyl)biphenyl-2,2′-diol 的抑制活性最高，为72%。 第三章综述了厚朴的化学成分及药理活性两个方面的研究进展。 The dissertation consists of three chapters. The first chapter is about the study on the constituents with α-glycosidase inhibitory activity from Magnolia officinalis. The EtOH extracts of M. officinalis Rehd. et Wils showed good inhibitory activity against α-Glucosidase. In order to determine the active compounds, bio-assay was used to guide the isolation. Six known alkaloids were isolated by solvent extraction and repeated silica gel column chromatography, and their structures were identified as liriodenine, anonaine, roemerine, lysicamine, magnoflorine and magnocurarine by spectroscopic methods. The inhibitory activity against α-Glucosidase of these alkaloids was measured with alvine screening model of α-glucosidase. Among them, lysicamine and liriodenine have the best inhibitory activity at 60% and 62%, respectively. The other four alkaloids have close inhibitory activity, from 46% to 51%. The second chapter is about the derivation of magnolol and the inhibitory a ctivity of the derivatives. Seven derivatives of magnolol were prepared by Manni-ch reaction, epoxidation followed by ring-opening reaction. Biological activity as say indicated the inhibitory activity was related to substituting groups. Among them, 5,5′-diallyl-3-((bis(2-hydroxyethyl)amino)methyl)biphenyl-2,2′-diol had the highest activity at 72%. The third chapter is a review on the progress of M. officinalis including chemical constituents and pharmacological activity.

吲哚生物碱Desbromoarborescidine A 的全合成及烯胺C-亚磺酰化反应研究

从新几内亚核桃木的树皮中分离得到的吲哚类喹诺里西定生物碱10-Desbromoarborescidine A，因发现其具有阻滞钙离子通道的活性而倍受关注。10-Desbromoarborescidine A由A、B、C、D四个环组成，只有一个手性中心，是吲哚生物碱中结构较简单的一种，常作为此类生物碱全合成方法的模型化合物。但迄今为止，能高效而简便的实现手性10-Desbromoarborescidine A不对称全合成方法线路不多，大多数以不对称诱导的方式建立其手性中心，手性催化的方式仅有一例金属催化。从逆合成分析可知，Desbromoarborescidine A的全合成可以通过亚胺不对称催化还原进行关键的手性中心构建，而本课题组在之前的研究中通过手性有机小分子催化剂的发展，已将三氯硅烷氢转移还原亚胺发展成了一类简便实用、高效、高对映选择性并具有优良底物适应范围的不对称催化反应，我们希望以这一反应作为关键手段，发展一条Desbromoarborescidine A及其类似物不对称合成新路线。 根据我们设计的新路线，首先成功合成了其关键中间体，然后我们进行了关键的不对称催化尝试。用本实验室已有的高性能有机小分子催化剂虽得到了较好的对应选择性，但是产率很低。同时，为了验证整条线路的可行性，我们也用消旋的中间体进行拉通线路的尝试。但不幸的是，在脱除保护基时遇到了很大困难。尝试换不同的保护基，或改变脱保护基的顺序，都未能成功合成目标产物。究其原因可能是由于吲哚的特殊性造成的，吲哚类亚胺与常规的芳香亚胺有较大的差异，其NH基团无论保护还是不保护，对与其2位相联接的C=N双键均有很大的影响，导致其不对称催化还原难以进行。另外，由于所设计的还原产物含有处在吲哚苄位的胺基，稳定性较差，造成保护基脱除困难。 烯胺C-亚磺酰化反应是本课题组最近发现的一个新反应，之前未见文献报道。本研究对该反应进行了反应条件优化和底物扩展，发现带Cbz，Ac，COt-Bu，CO2Et，Bz等保护基的一系列环状和非环状烯胺在亚磺酸钠、DMAc和MeSiCl3的共同作用下能高效高产率生成β-胺基烯基亚砜类新化合物，为合成多官能团化的烯基亚砜新化合物提供了一条简便实用的途径。 The main constituent of Dracontomelum mangiferum B1, indoloquinolizidine alkaloid 10-Desbromoarborescidine A, has drawn great attention due to its calcium channel blocking activity. Its molecular structure is relatively simple compared with the other alkaloids of the same type, which has only one chiral center, albeit with four cycles A, B, C, and D. This compound is often used as a model target for exploring different strategies for the total synthesis of indole alkaloids. Nevertheless, so far there still lack practical and highly efficient methods for the asymmetric total synthesis of 10-Desbromoarborescidine A. Most of the current available methods rely on stoichiometric asymmetric synthesis for the construction of the chiral center. There is only one example reporting utilization of asymmetric catalysis, but with transition metal complex as the catalyst. Our retrosynthetic analysis shows that catalytic asymmetric reduction of imine could be used as the key step for the construction of the chiral center of Desbromoarborescidine A. Since in the previous studies our group has developed the asymmetric reduction of imines by trichlorosilane into a practical and highly efficient and enantioselective method using newly designed chiral organocatalysts, we hope to apply this method to develop a novel synthetic route for the total synthesis of Desbromoarborescidine A and its analogues in this study. According to the newly designed synthetic route, we first accomplished the synthesis of the key intermediates which was then examined for the critical asymmetric catalysis. The asymmetric reduction using the highly efficient organocatalysts, developed in our lab afforded high ee but poor yield. We tried different reaction conditions to improve the yield, but failed to get any good results. Simultaneously, to vertify the feasibility of the synthetic route we designed, we also tired to go through the route toward the racemic synthesis of Desbromoarborescidine A. But unfortunately, protection and deprotection proved to be big hurdles. All the different protection groups and different sequences of protection and deprotection we tried failed to get us through the designed synthetic sequence and furnish the final product. Most likely, the indole part is the culprit behind the failures.The NH group of the indole, no matter protected or not, may impact the catalytic asymmetric reduction of C-N double bond connected with 2-C. Additionally, the reduction product we designed contains an amino group in the β-position of the indole, which may cause problems due to its instability. C-sulfenylation of enamines is a novel reaction discovered recently by our group, which has not been seen before in the literature. In this study, optimization of the reaction conditions and exploration of the substrate scope were further undertaken for this reaction, which reveal that a series of enamines with N-Cbz, Ac, COt-Bu, CO2Et protection groups could all undergo smooth C-sulfinylations with the comined use of sodium benzene sulphinate, DAMc and MeSiCl3, efficiently furnishing the β-amino vinylsulfoxide products in high yield, affording a practical and highly efficient methods for synthesis of functional vinylsulfoxides.

Early effects of low dose C-12(6+) ion or X-ray irradiation on human peripheral blood lymphocytes

The aim of this study was to estimate the acute effects of low dose C-12(6+) ions or X-ray radiation on human immune function. The human peripheral blood lymphocytes (HPBL) of seven healthy donors were exposed to 0.05 Gy C-12(6+) ions or X-ray radiation and cell responses were measured at 24 h after exposure. The cytotoxic activities of HPBL were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT); the percentages of T and NK cells subsets were detected by flow cytometry; mRNA expression of interleukin (IL)-2, tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma were examined by real time quantitative RT-PCR (qRT-PCR); and these cytokines protein levels in supematant of cultured cells were assayed by enzyme-linked immunosorbent assays (ELISA). The results showed that the cytotoxic activity of HPBL, mRNA expression of IL-2, IFN-gamma and TNF-alpha in HPBL and their protein levels in supernatant were significantly increased at 24 h after exposure to 0.05 Gy C-12(6+) ions radiation and the effects were stronger than observed for X-ray exposure. However, there was no significant change in the percentage of T and NK cells subsets of HPBL. These results suggested that 0.05 Gy high linear energy transfer (LET) C-12(6+) radiation was a more effective approach to host immune enhancement than that of low LET X-ray. We conclude that cytokines production might be used as sensitive indicators of acute response to LDL (C) 2009 COSPAR. Published by Elsevier Ltd. All rights reserved.