3-正丁基苯酞体内药动学及脑内转运机理的研究
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摘要
3-正丁基苯酞(3-n-butylphthalide，简称NBP)是从芹菜籽挥发油中分离得到的抗脑缺血的有效成分，经化学合成后用于临床试验的新药。本文旨在研究NBP在体内的药动学过程及组织分布情况，考察其在肠粘膜的吸收性质及肝脏的首过效应，从而阐明影响其口服吸收的因素；研究NBP向靶器官—脑组织转运的机制，为合理给药途径的提出及剂型的设计提供理论依据；通过考察NBP在鼻粘膜的吸收性质及对鼻纤毛的毒性，论证NBP鼻腔给药的可行性；并通过研究NBP脂质体经鼻给药的生物利用度及向脑内转运的途径对实验设想予以证实。
首先，建立了NBP在家兔和大鼠体内的测定方法，采用HPLC法以荧光检测器进行检测。色谱条件为：以0.05 mol/L醋酸钠(用醋酸调pH4.5)-乙腈(40：60)为流动相，采用Selectosil C_(18)色谱柱，检测波长为λ_(ex)=280 nm，λ_(em)=304 nm，以布洛芬为内标，样品经乙醚提取处理测定。该法专属性强，灵敏度高，适于NBP体内药动学研究。
分别以家兔和大鼠为模型，考察了NBP在不同动物体内的药动学过程：NBP在家兔和大鼠体内均呈线性动力学过程，符合双隔室开放模型。静注给药后从体内消除迅速：不同剂量组NBP在家兔体内的T_(1/2α)为10.3～11.3 min，T_(1/2β)为96～109min：在大鼠体内的T_(1/2α)为4.7～9min，T_(1/2β)为68～125 min。NBP分别以羟丙基-β-环糊精(Hydroxypropyl-β-cyclodextrin，简称HPCD)包合物和脂质体静注给药，并不影响NBP在大鼠体内的动力学过程。NBP口服给药后，吸收迅速，在家兔和大鼠体内达峰时间(T_(max))约为30 min。家兔口服生物利用度为4.7±1.7％；于大鼠口服给药后，NBP脂质体的生物利用度(30.3±9.6％)与NBP溶液(50％1，2-丙二醇溶液)相比(26.3±10.3％)无显著性差异(P＞0.05)；NBP-HPCD包合物的生物利用度显著降低(15.0±3.7％)。
采用同位素示踪技术考察了[~3H]NBP在大鼠体内的分布，NBP主要分布于肝、肾、肺、脑、脾等组织，各组织的浓度均呈一定的剂量依赖性，NBP在大脑、嗅球、小脑、脑室、脑干的浓度间无显著性差异(P＞0.05)。
以大鼠在体肠灌流技术(In-situ single-pass perfusion)考察了NBP在肠粘膜的吸收性质，NBP在大鼠十二指肠、空肠、回肠和结肠的表观吸收系数分别为0.0381±0.0103，0.0314±0.0041，0.0277±0.0060和0.0574±0.0050 ml/min/cm(灌流液中NBP浓度为10 μg/ml)，NBP浓度对肠段的表观吸收系数无明显影响(P＞0.05)。表明NBP在各肠段的吸收均呈被动扩散机制，也说明肠粘膜吸收过程非NBP口服生物利用度低的主要影响因素。灌流液中HPCD的加入对NBP在大鼠肠粘膜的吸收具有较强的抑制作用，其抑制程度随HPCD用量的增加而显著增强。而NBP包
沈阳药科大学博士学位论文
摘要
封于脂质体中后对NBP在大鼠肠粘膜的吸收无显著性影响。说明NBP一HPCD包合
物于大鼠口服生物利用度低是由HPCD的抑制作用所引起石
采用离体大鼠肝灌流技术测得不同浓度NBP(2，5和20林g/inl)在肝脏的清
除率分别为6.31士0.45，3.14士0.15，2.71士0.48 ml/m in;HPCD对NBP在离体大
鼠肝脏的消除有抑制作用;NBP脂质体对NBP在大鼠离体肝脏的消除无显著性影
响;尼莫地平对NBP的消除无显著性影响。原代大鼠肝细胞对护H]NBP的摄取实
验表明，NBP能迅速进入离体肝细胞中并达到稳态，且细胞中NBP浓度比培养介
质中高约4倍，细胞的摄取过程无自身浓度抑制作用，呈能量非依赖性过程，说
明NBP进入肝细胞为被动扩散机制。由以上实验结果表明NBP在大鼠肝脏具有较
强的首过效应，由此可解释NBP口服生物利用度低和NBP在肝脏中浓度高的现象。
离体脉络丛摄取实验表明，脉络丛对NBP的摄取无自身浓度抑制作用，呈能
量非依赖性转运过程，且NBP的T/M值均较高(1 4.巧士1 .18)。说明NBP在大鼠
脉络丛中的转运主要为被动扩散机制，NBP易于透过脑屏障。尼莫地平和桂利嗦
对NBP在离体大鼠脉络丛的转运方式无显著性影响(P>0.05)。
NBP在大鼠鼻粘膜吸收迅速，用药后5 min即可吸收55%以上;但对大鼠鼻
粘膜具有较强的纤毛毒性，NBP经HPCD包合后可显著降低NBP的纤毛毒性，但
对NBP在鼻粘膜的吸收有一定抑制作用。NBP包封于脂质体中后不但显著降低了
纤毛毒性，且对NBP在鼻粘膜的吸收无显著性影响。说明脂质体可作为NBP鼻腔
给药的剂型之一。
采用超声一高压乳匀法制备了NBP脂质体。磷脂的种类和用量、载药量以及胆
固醇用量对脂质体的包封率均有一定的影响。以合成DPPC为磷脂膜的脂质体包
封率较高且稳定;随载药量的增大包封率显著降低;制备工艺对脂质体粒径有显
著性影响。以超声一高压乳匀法制得的脂质体有效粒径为88.snln，聚分散度为
0 .1 84。
NBP脂质体于家兔经鼻给药后，迅速吸收进入体循环，2一5 min即可达最大血
药浓度，体内药时过程与静注给药相似，其绝对生物利用度为88.7%。于大鼠经鼻
给药后，2 min即可在脑组织中达到较高的浓度，尤其以脑组织的嗅球部位浓度最
高。30一60 min NBP在脑组织中的浓度显著高于静注给药。说明NBP经鼻给药后
进入脑组织的途径有两条:a)嗅粘膜上皮细胞途
Zhao Chunshun (Major: Pharmaceutics) Advisors: Prof. Zhang Ruhua and Associate Prof. He Zhonggui
3-n-Butylphthalide (NBP), a novel cerebral antiischemic agent, was isolated and identified from several plants including celery oil. At present, NBP was synthesized and approved for Phase IV trials in treatment of stroke by the State Drug Administration of China.
The main objectives of this research were to study the pharmacokinetics, absolute bioavailability, disposition, intestinal absorption and first-pass effect of NBP in rabbits and rats, to investigate the mechanism of transport of NBP in the blood-cerebrospinal fluid barrier, to research nasal mucosa absorption and nasal ciliotoxicity of NBP in rats, and to develop reasonable delivery system of NBP used in intranasal administration. We also determined the uptake of NBP into the brain after intravenous and intranasal administration in rats.
A rapid, sensitive and specific RP-HPLC method was developed for the determination of NBP in rabbits or rats plasma in combination with fluorescence detection at an excitation wavelength of 280 nm and an emission wavelength of 304 nm. Ibuprofen was used as internal standard. Plasma samples were extracted with diethyl ether under acidic conditions. After evaporation of the organic phase, the extract was dissolved in mobile phase and injected into the chromatograph with C18 column and a mobile phase of 0.05mol/L sodium acetate buffer (pH 4.5)-acetonitrile (400:600). The achieved limit of quantification of 0.0212 ug/ml is sufficient to study the pharmacokinetics of NBP in rabbits or rats.
The pharmacokinetic investigation were performed on rabbits and rats. NBP exhibited linear pharmacokinetics over the dose range tested (l-10mg/kg) in rabbits and rats after intravenous administration. NBP was rapidly eliminated from the plasma. The mean elimination (T1/2a,10.3-11.3 min and T1/2B, 96-109 min in rabbits; T1/2a, 4.7-9 min and T1/2B, 68-125 min in rats), total plasma clearance, and apparent volume values were independent of doses. The dosage forms had no effect on the pharmacokinetic of NBP in rats. After oral administration to rabbits and rats, NBP reached the peak plasma concentration at a time ranging between 10 to 45 min. Oral bioavailability was lower in
rabbits (4.7±1.7%) than in rats (26.3±10.3%) for NBP solution. NBP-HPCD inclusion reduced the bioavailability (15.0±3.7%) of NBP in rats.
The distribution of [3H]NBP in tissues was determined at 10, 30, 60, and 90 min after intravenous administration of a single 1, 5, and 10 mg/kg dose to rats by liquid scintillation counter. The highest concentration was found in the liver, and the concentration decreased in the order of liver, kidney, lung, brain, heart and spleen. NBP in tissues is dose-dependent. NBP concentration was not significantly different among cerebrum, cerebellum, olfactory bulb, cerebral ventricle and brain stem.
The intestinal transport of NBP was performed by applying single-pass perfusion techniques in situ rats. NBP is a high permeability drug in rat intestine and colon. The apparent permeability, Papp, of NBP was independent of both intestinal region and concentration of NBP present in the perfusion solution. The Papp for NBP in the colon is higher than that in various intestinal regions. The Papp for NBP decreased when HPCD is co-perfused in the intestine and colon. The absorption inhibitor effect of HPCD was concentration dependent and it was supposed that the concentration of free NBP is too low in the perfusion solution when HPCD was added in.
The hepatic clearance of NBP during isolated rat liver perfusion were 6.31±0.45, 3.14±0.15, and 2.71±0.48 ml/min for 2, 5, and 10 ug/ml, respectively. The elimination of NBP was inhibited when HPCD is co-perfused in the isolated liver. Liposomes didn't influence the elimination of NBP from the isolated rat liver. Uptake of [3H]NBP by rat primary hepatocytes was rapid and a steady-state was achieved in 15 seconds. The accumulation of [3H]NBP in hepatocytes wasn't inhibited by unlabeled NBP,
首先，建立了NBP在家兔和大鼠体内的测定方法，采用HPLC法以荧光检测器进行检测。色谱条件为：以0.05 mol/L醋酸钠(用醋酸调pH4.5)-乙腈(40：60)为流动相，采用Selectosil C_(18)色谱柱，检测波长为λ_(ex)=280 nm，λ_(em)=304 nm，以布洛芬为内标，样品经乙醚提取处理测定。该法专属性强，灵敏度高，适于NBP体内药动学研究。
分别以家兔和大鼠为模型，考察了NBP在不同动物体内的药动学过程：NBP在家兔和大鼠体内均呈线性动力学过程，符合双隔室开放模型。静注给药后从体内消除迅速：不同剂量组NBP在家兔体内的T_(1/2α)为10.3～11.3 min，T_(1/2β)为96～109min：在大鼠体内的T_(1/2α)为4.7～9min，T_(1/2β)为68～125 min。NBP分别以羟丙基-β-环糊精(Hydroxypropyl-β-cyclodextrin，简称HPCD)包合物和脂质体静注给药，并不影响NBP在大鼠体内的动力学过程。NBP口服给药后，吸收迅速，在家兔和大鼠体内达峰时间(T_(max))约为30 min。家兔口服生物利用度为4.7±1.7％；于大鼠口服给药后，NBP脂质体的生物利用度(30.3±9.6％)与NBP溶液(50％1，2-丙二醇溶液)相比(26.3±10.3％)无显著性差异(P＞0.05)；NBP-HPCD包合物的生物利用度显著降低(15.0±3.7％)。
采用同位素示踪技术考察了[~3H]NBP在大鼠体内的分布，NBP主要分布于肝、肾、肺、脑、脾等组织，各组织的浓度均呈一定的剂量依赖性，NBP在大脑、嗅球、小脑、脑室、脑干的浓度间无显著性差异(P＞0.05)。
以大鼠在体肠灌流技术(In-situ single-pass perfusion)考察了NBP在肠粘膜的吸收性质，NBP在大鼠十二指肠、空肠、回肠和结肠的表观吸收系数分别为0.0381±0.0103，0.0314±0.0041，0.0277±0.0060和0.0574±0.0050 ml/min/cm(灌流液中NBP浓度为10 μg/ml)，NBP浓度对肠段的表观吸收系数无明显影响(P＞0.05)。表明NBP在各肠段的吸收均呈被动扩散机制，也说明肠粘膜吸收过程非NBP口服生物利用度低的主要影响因素。灌流液中HPCD的加入对NBP在大鼠肠粘膜的吸收具有较强的抑制作用，其抑制程度随HPCD用量的增加而显著增强。而NBP包
沈阳药科大学博士学位论文
摘要
封于脂质体中后对NBP在大鼠肠粘膜的吸收无显著性影响。说明NBP一HPCD包合
物于大鼠口服生物利用度低是由HPCD的抑制作用所引起石
采用离体大鼠肝灌流技术测得不同浓度NBP(2，5和20林g/inl)在肝脏的清
除率分别为6.31士0.45，3.14士0.15，2.71士0.48 ml/m in;HPCD对NBP在离体大
鼠肝脏的消除有抑制作用;NBP脂质体对NBP在大鼠离体肝脏的消除无显著性影
响;尼莫地平对NBP的消除无显著性影响。原代大鼠肝细胞对护H]NBP的摄取实
验表明，NBP能迅速进入离体肝细胞中并达到稳态，且细胞中NBP浓度比培养介
质中高约4倍，细胞的摄取过程无自身浓度抑制作用，呈能量非依赖性过程，说
明NBP进入肝细胞为被动扩散机制。由以上实验结果表明NBP在大鼠肝脏具有较
强的首过效应，由此可解释NBP口服生物利用度低和NBP在肝脏中浓度高的现象。
离体脉络丛摄取实验表明，脉络丛对NBP的摄取无自身浓度抑制作用，呈能
量非依赖性转运过程，且NBP的T/M值均较高(1 4.巧士1 .18)。说明NBP在大鼠
脉络丛中的转运主要为被动扩散机制，NBP易于透过脑屏障。尼莫地平和桂利嗦
对NBP在离体大鼠脉络丛的转运方式无显著性影响(P>0.05)。
NBP在大鼠鼻粘膜吸收迅速，用药后5 min即可吸收55%以上;但对大鼠鼻
粘膜具有较强的纤毛毒性，NBP经HPCD包合后可显著降低NBP的纤毛毒性，但
对NBP在鼻粘膜的吸收有一定抑制作用。NBP包封于脂质体中后不但显著降低了
纤毛毒性，且对NBP在鼻粘膜的吸收无显著性影响。说明脂质体可作为NBP鼻腔
给药的剂型之一。
采用超声一高压乳匀法制备了NBP脂质体。磷脂的种类和用量、载药量以及胆
固醇用量对脂质体的包封率均有一定的影响。以合成DPPC为磷脂膜的脂质体包
封率较高且稳定;随载药量的增大包封率显著降低;制备工艺对脂质体粒径有显
著性影响。以超声一高压乳匀法制得的脂质体有效粒径为88.snln，聚分散度为
0 .1 84。
NBP脂质体于家兔经鼻给药后，迅速吸收进入体循环，2一5 min即可达最大血
药浓度，体内药时过程与静注给药相似，其绝对生物利用度为88.7%。于大鼠经鼻
给药后，2 min即可在脑组织中达到较高的浓度，尤其以脑组织的嗅球部位浓度最
高。30一60 min NBP在脑组织中的浓度显著高于静注给药。说明NBP经鼻给药后
进入脑组织的途径有两条:a)嗅粘膜上皮细胞途
Zhao Chunshun (Major: Pharmaceutics) Advisors: Prof. Zhang Ruhua and Associate Prof. He Zhonggui
3-n-Butylphthalide (NBP), a novel cerebral antiischemic agent, was isolated and identified from several plants including celery oil. At present, NBP was synthesized and approved for Phase IV trials in treatment of stroke by the State Drug Administration of China.
The main objectives of this research were to study the pharmacokinetics, absolute bioavailability, disposition, intestinal absorption and first-pass effect of NBP in rabbits and rats, to investigate the mechanism of transport of NBP in the blood-cerebrospinal fluid barrier, to research nasal mucosa absorption and nasal ciliotoxicity of NBP in rats, and to develop reasonable delivery system of NBP used in intranasal administration. We also determined the uptake of NBP into the brain after intravenous and intranasal administration in rats.
A rapid, sensitive and specific RP-HPLC method was developed for the determination of NBP in rabbits or rats plasma in combination with fluorescence detection at an excitation wavelength of 280 nm and an emission wavelength of 304 nm. Ibuprofen was used as internal standard. Plasma samples were extracted with diethyl ether under acidic conditions. After evaporation of the organic phase, the extract was dissolved in mobile phase and injected into the chromatograph with C18 column and a mobile phase of 0.05mol/L sodium acetate buffer (pH 4.5)-acetonitrile (400:600). The achieved limit of quantification of 0.0212 ug/ml is sufficient to study the pharmacokinetics of NBP in rabbits or rats.
The pharmacokinetic investigation were performed on rabbits and rats. NBP exhibited linear pharmacokinetics over the dose range tested (l-10mg/kg) in rabbits and rats after intravenous administration. NBP was rapidly eliminated from the plasma. The mean elimination (T1/2a,10.3-11.3 min and T1/2B, 96-109 min in rabbits; T1/2a, 4.7-9 min and T1/2B, 68-125 min in rats), total plasma clearance, and apparent volume values were independent of doses. The dosage forms had no effect on the pharmacokinetic of NBP in rats. After oral administration to rabbits and rats, NBP reached the peak plasma concentration at a time ranging between 10 to 45 min. Oral bioavailability was lower in
rabbits (4.7±1.7%) than in rats (26.3±10.3%) for NBP solution. NBP-HPCD inclusion reduced the bioavailability (15.0±3.7%) of NBP in rats.
The distribution of [3H]NBP in tissues was determined at 10, 30, 60, and 90 min after intravenous administration of a single 1, 5, and 10 mg/kg dose to rats by liquid scintillation counter. The highest concentration was found in the liver, and the concentration decreased in the order of liver, kidney, lung, brain, heart and spleen. NBP in tissues is dose-dependent. NBP concentration was not significantly different among cerebrum, cerebellum, olfactory bulb, cerebral ventricle and brain stem.
The intestinal transport of NBP was performed by applying single-pass perfusion techniques in situ rats. NBP is a high permeability drug in rat intestine and colon. The apparent permeability, Papp, of NBP was independent of both intestinal region and concentration of NBP present in the perfusion solution. The Papp for NBP in the colon is higher than that in various intestinal regions. The Papp for NBP decreased when HPCD is co-perfused in the intestine and colon. The absorption inhibitor effect of HPCD was concentration dependent and it was supposed that the concentration of free NBP is too low in the perfusion solution when HPCD was added in.
The hepatic clearance of NBP during isolated rat liver perfusion were 6.31±0.45, 3.14±0.15, and 2.71±0.48 ml/min for 2, 5, and 10 ug/ml, respectively. The elimination of NBP was inhibited when HPCD is co-perfused in the isolated liver. Liposomes didn't influence the elimination of NBP from the isolated rat liver. Uptake of [3H]NBP by rat primary hepatocytes was rapid and a steady-state was achieved in 15 seconds. The accumulation of [3H]NBP in hepatocytes wasn't inhibited by unlabeled NBP,
引文
[1]方洪巨；杨竣山；胡秋．挥发油成分的研究／V．芹菜子挥发油的化学成分研究．药物分析杂志，1984，4(4)：205—208．
[2]杨峻山；陈玉武．芹菜抗惊有效成分的分离和鉴定．药学通报，1984，19(11)：30—31．
[3] Bjeldanes, Leonard F.; Kim, In-Suk. Sedative activity of celery oil constituents. J. Food Sci.,1978,43(1):143-144.
[4] Rao, L. Jagan Mohan; Nagalakshmi, S.; Naik, J. Pura; Shankaracharya, N. B. Studies on
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[5] Ludwiczuk, A.; Najda, A.; Wolski, T.; Baj, T. Chromatographic determination of the content and the composition of extracts and essential oils from the fruits of three varieties of stalk celery (Apium graveolens L. var. dulce mill. pers.). Journal of Planar Chromatography-Modern TLC,
2001,14(6):400-404.
[6]王普善；高宣亮；福山爱．中药川芎的化学成分研究—五种内酯类化合物．中草药，1985，16(3)：41．
[7] Li, Hong-Xia; Ding, Ming-Yu; Yu, Jian-Yuan. Separation and identification of the phthalic anhydride derivatives of Ligusticum Chuanxiong hort by GC-MS, TLC, HPLC-DAD, and HPLC-MS. Journal of Chromatographic Science, 2002,40(3): 156-161.
[8]王文祥；顾明；蒋小岗．川芎化学成分研究．中草药，2002,33(1)：4—5．
[9]罗永明；张金海；潘家洁等．特产中药茶芎化学成分的研究．中国药学杂志，1994,29(12)：714—715．
[10]罗永明；潘家祜；丁科平．茶芎挥发油中抗惊有效成分的分离和鉴定．中草药，1996,27(8)：456-457．
[11]Gijbels, M.J.M.; Bos, R.; Scheffer, J.J.C.; Svendsen, A. Baerheim. Phthalides in roots of
Opopanax chironium. Planta Med., 1983,47(1):3-6.
[12]Lin, Long-Ze; He, Xian-Guo; Lian Li-Zhi; King, Wayne; Elliott, Jerry. Liqiud chromatographic-
electroshpray mass spectrometric study of the phthalides of Angelica sinensis and chemical
changes of Z-ligustilide. J. Chromatogr., A, 1998,810(1+2):71-79.
[13]席与；孙明杰；李惟明．藁本化学成分的研究．中草药，1987，18(2)：6-7．
[14]黄远征；溥发鼎．几种藁本属植物挥发油化学成分的分析．药物分析杂志，1989,9(3)：147-151．
[15]张金兰；何秀峰；周志华．藁本中5种成分的高效液相色谱法测定．药学学报，1996,31(8)：622—625．
[16]江滨；韦群辉；许晓峰．滇藁本精油中镇静成分的分离与鉴定．中国民族民间医药杂志，1998，4：34-35
[17]Kitayama, Takashi. Microbial asymmetric synthesis of 3-alkylphthalide derivatives. Tetrahedron:Asymmetry, 1997,8(22):3765-3774.
[18]Martinez, M. Montserrat; Onega, M. Gabriela; Fe Tellado, M.; Seijas, Julio A.; Vazquez-Tato, M.
Pilar. 1,6-Conjugate addition to o-vinylphenyloxazolines. Synthesis of chuanxinol and 3-n-butylphthalide. Tetrahedron, 1997,53(41):14127-14130.
[19]李仲辉，陈孝康，张晓南等．降压新药(±)—芹菜甲素1a～1d合成方法研究．四川教育学院学报，2000，16(1—2)：90-91．
[20]阎福林，李绍白，孙祥德．3—烃基苯酞类化合物的合成研究．郑州工业大学学报，1999,20(2)：48-50．
[21]Kawasaki, Taishi; Saito, Shinichi; Yamamoto, Yoshinori. Synthesis of Phthalides and 3,4-Dihydroisocoumarins Using the Palladium-Catalyzed Intramolecular Benzannulation Strategy. Journal of Organic Chemistry, 2002, 67(8):2653-2658.
[22]Guo, Zongru; Chu, Fengming; Zhang, Juntian; Yang, Guangzhong; Xu, Bailing; Niu, Xinyi; Ren, Zhihong; Lestage, Pierre; Renard, Pierre. Preparation of new substituted phthalides as anticonvulsants. Peop. Rep. China, PCT Int. Appl.WO 2002000638 A2 3 Jan 2002.
[23]陈孝康，李仲辉，陶玲，王守文．“一锅法”合成(±)-3-烷基苯酞．合成化学，1999，7(3)：292—294
[24]于澍仁，尤胜权．芹菜甲素和乙素的抗惊厥作用．药学学报，1984，19(8)：566-570．
[25]董高翔；冯亦璞．丁基苯酞及其光学异构体的抗惊厥作用．中国药理学通报，1999，15(1)：88-89．
[26]冯亦璞．缺血性脑卒中的病理生理及药物治疗现状．药学学报，1999,34(1)：72—78．
[27]冯亦璞；胡盾；张丽英．丁基苯酞对小鼠全脑缺血的保护作用．药学学报，1995,30(10)：741-744．
[28]熊杰；冯亦璞．丁基苯酞对线粒体呼吸链复合酶活性的影响．药学学报，1999,34(4)：241-245
[29]Yan CH, Feng YP, Zhang JT. Effects of dl-3-n-butylphthalide on region al cerebral blood flow in right middle cerebral artery occlusion rats, Acta Pharmacol Sin, 1998,19(2):117-120.
[30]种兆忠；冯亦璞．丁基苯酞改善大鼠实验性蛛网膜下腔出血后局部脑血流．中国药理学报，1999,20(6)：509-512．
[31]刘小光；冯亦璞．丁基苯酞对局部脑缺血大鼠行为和病理改变的保护作用．药学学报，1995，30(12)：896-903．
[32]Deng, Wenbin; Feng, Yipu. Effect of dl-3-n-butylphthalide on brain edema in rats subjected to focal cerebral ischemia. Chin. Med. Sci. J., 1997,12(2):102-106.
[33]胡盾；张丽英；冯亦璞．丁基苯酞对局部脑缺血大鼠记忆障碍的影响．中国药理学与毒理学杂志，1997，11(1)：14-16．
[34]张丽英；冯亦璞．丁基苯酞对脑卒中型自发型高血压大鼠寿命及卒中后神经症状的影响．药学学报，1996,31(1)：18-23．
[35]阎超华；张均田；冯亦璞．丁基苯酞对氯化钾及N-甲基-D-门冬氨酸诱导的大鼠皮质神经
细胞损伤的保护作用．药学学报，1997,32(5)：340-346．
[36]阎超华；冯亦璞．丁基苯酞对低糖低氧诱导的大鼠皮层神经细胞损伤的保护作用．药学学
报，1998，33(7)：486—492．
[37]董高翔；冯亦璞．丁基苯酞抑制低氧低糖诱导的大鼠皮质神经细胞凋亡．药学学报，1999，34(3)：176-180．
[38]刘小光；冯亦璞．丁基苯酞对氯化钾和去甲肾上腺素引起的离体大鼠尾动脉环收缩的影响．
中国药理学与毒理学杂志，1996，10(2)：113—115．
[39]熊杰；冯亦璞．丁基苯酞对低糖低氧引起神经细胞内钙升高的作用．药学学报，1999,34(12)：893-897．
[40]种兆忠；冯亦璞．丁基苯肽对大脑中动脉阻断后皮层组织中花生四烯酸释放及磷脂酶A_2
基因表达的影响．药学学报，2000,35(8)：561～565．
[41]熊杰；冯亦璞．丁基苯酞对局灶性脑缺血过程中线粒体损伤的保护作用．药学学报，2000，
35(6)：408—412．
[42]董高翔；冯亦璞．丁基苯酞对局部脑缺血再灌注大鼠脑线粒体ATPase，抗氧化酶活性和脂
质过氧化的影响．中国医学科学院学报，2002,24(1)：93—97．
[43]种兆忠；冯亦璞．丁基苯酞对大鼠皮层神经元损伤后谷氨酸和5-羟色胺释放的影响．中国
药学杂志．1999,34(9)：589—591．
[44]胡盾；黄新祥；冯亦璞．丁基苯酞对全脑缺血大鼠的纹状体细胞外液嘌呤类代谢物含量的
影响．药学学报，1996,31(1)：13-17．
[45]种兆忠；冯亦璞丁基苯酞对大鼠局灶性脑缺血和重灌后脑内TXB-2和6-keto-PGF_(1a)含量
的影响中国药理学报1997，8(6)：505-508．
[46]Peng SH, Zhou TH. The study on in vitro metabolism of butylphthalide by gas chromatography-mass spectrometry. Chin Chem Lett, 1995, 6:55.
[47]彭仕华；周同惠．丁基苯酞的体内代谢转化研究．药学学报，1996,31(10)：780-784．
[48]王春华；冯亦璞；吴元鎏．丁基苯酞在大鼠中代谢产物的研究．药学学报，1997,32(9)：641-646．
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[2]杨峻山；陈玉武．芹菜抗惊有效成分的分离和鉴定．药学通报，1984，19(11)：30—31．
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[4] Rao, L. Jagan Mohan; Nagalakshmi, S.; Naik, J. Pura; Shankaracharya, N. B. Studies on
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[8]王文祥；顾明；蒋小岗．川芎化学成分研究．中草药，2002,33(1)：4—5．
[9]罗永明；张金海；潘家洁等．特产中药茶芎化学成分的研究．中国药学杂志，1994,29(12)：714—715．
[10]罗永明；潘家祜；丁科平．茶芎挥发油中抗惊有效成分的分离和鉴定．中草药，1996,27(8)：456-457．
[11]Gijbels, M.J.M.; Bos, R.; Scheffer, J.J.C.; Svendsen, A. Baerheim. Phthalides in roots of
Opopanax chironium. Planta Med., 1983,47(1):3-6.
[12]Lin, Long-Ze; He, Xian-Guo; Lian Li-Zhi; King, Wayne; Elliott, Jerry. Liqiud chromatographic-
electroshpray mass spectrometric study of the phthalides of Angelica sinensis and chemical
changes of Z-ligustilide. J. Chromatogr., A, 1998,810(1+2):71-79.
[13]席与；孙明杰；李惟明．藁本化学成分的研究．中草药，1987，18(2)：6-7．
[14]黄远征；溥发鼎．几种藁本属植物挥发油化学成分的分析．药物分析杂志，1989,9(3)：147-151．
[15]张金兰；何秀峰；周志华．藁本中5种成分的高效液相色谱法测定．药学学报，1996,31(8)：622—625．
[16]江滨；韦群辉；许晓峰．滇藁本精油中镇静成分的分离与鉴定．中国民族民间医药杂志，1998，4：34-35
[17]Kitayama, Takashi. Microbial asymmetric synthesis of 3-alkylphthalide derivatives. Tetrahedron:Asymmetry, 1997,8(22):3765-3774.
[18]Martinez, M. Montserrat; Onega, M. Gabriela; Fe Tellado, M.; Seijas, Julio A.; Vazquez-Tato, M.
Pilar. 1,6-Conjugate addition to o-vinylphenyloxazolines. Synthesis of chuanxinol and 3-n-butylphthalide. Tetrahedron, 1997,53(41):14127-14130.
[19]李仲辉，陈孝康，张晓南等．降压新药(±)—芹菜甲素1a～1d合成方法研究．四川教育学院学报，2000，16(1—2)：90-91．
[20]阎福林，李绍白，孙祥德．3—烃基苯酞类化合物的合成研究．郑州工业大学学报，1999,20(2)：48-50．
[21]Kawasaki, Taishi; Saito, Shinichi; Yamamoto, Yoshinori. Synthesis of Phthalides and 3,4-Dihydroisocoumarins Using the Palladium-Catalyzed Intramolecular Benzannulation Strategy. Journal of Organic Chemistry, 2002, 67(8):2653-2658.
[22]Guo, Zongru; Chu, Fengming; Zhang, Juntian; Yang, Guangzhong; Xu, Bailing; Niu, Xinyi; Ren, Zhihong; Lestage, Pierre; Renard, Pierre. Preparation of new substituted phthalides as anticonvulsants. Peop. Rep. China, PCT Int. Appl.WO 2002000638 A2 3 Jan 2002.
[23]陈孝康，李仲辉，陶玲，王守文．“一锅法”合成(±)-3-烷基苯酞．合成化学，1999，7(3)：292—294
[24]于澍仁，尤胜权．芹菜甲素和乙素的抗惊厥作用．药学学报，1984，19(8)：566-570．
[25]董高翔；冯亦璞．丁基苯酞及其光学异构体的抗惊厥作用．中国药理学通报，1999，15(1)：88-89．
[26]冯亦璞．缺血性脑卒中的病理生理及药物治疗现状．药学学报，1999,34(1)：72—78．
[27]冯亦璞；胡盾；张丽英．丁基苯酞对小鼠全脑缺血的保护作用．药学学报，1995,30(10)：741-744．
[28]熊杰；冯亦璞．丁基苯酞对线粒体呼吸链复合酶活性的影响．药学学报，1999,34(4)：241-245
[29]Yan CH, Feng YP, Zhang JT. Effects of dl-3-n-butylphthalide on region al cerebral blood flow in right middle cerebral artery occlusion rats, Acta Pharmacol Sin, 1998,19(2):117-120.
[30]种兆忠；冯亦璞．丁基苯酞改善大鼠实验性蛛网膜下腔出血后局部脑血流．中国药理学报，1999,20(6)：509-512．
[31]刘小光；冯亦璞．丁基苯酞对局部脑缺血大鼠行为和病理改变的保护作用．药学学报，1995，30(12)：896-903．
[32]Deng, Wenbin; Feng, Yipu. Effect of dl-3-n-butylphthalide on brain edema in rats subjected to focal cerebral ischemia. Chin. Med. Sci. J., 1997,12(2):102-106.
[33]胡盾；张丽英；冯亦璞．丁基苯酞对局部脑缺血大鼠记忆障碍的影响．中国药理学与毒理学杂志，1997，11(1)：14-16．
[34]张丽英；冯亦璞．丁基苯酞对脑卒中型自发型高血压大鼠寿命及卒中后神经症状的影响．药学学报，1996,31(1)：18-23．
[35]阎超华；张均田；冯亦璞．丁基苯酞对氯化钾及N-甲基-D-门冬氨酸诱导的大鼠皮质神经
细胞损伤的保护作用．药学学报，1997,32(5)：340-346．
[36]阎超华；冯亦璞．丁基苯酞对低糖低氧诱导的大鼠皮层神经细胞损伤的保护作用．药学学
报，1998，33(7)：486—492．
[37]董高翔；冯亦璞．丁基苯酞抑制低氧低糖诱导的大鼠皮质神经细胞凋亡．药学学报，1999，34(3)：176-180．
[38]刘小光；冯亦璞．丁基苯酞对氯化钾和去甲肾上腺素引起的离体大鼠尾动脉环收缩的影响．
中国药理学与毒理学杂志，1996，10(2)：113—115．
[39]熊杰；冯亦璞．丁基苯酞对低糖低氧引起神经细胞内钙升高的作用．药学学报，1999,34(12)：893-897．
[40]种兆忠；冯亦璞．丁基苯肽对大脑中动脉阻断后皮层组织中花生四烯酸释放及磷脂酶A_2
基因表达的影响．药学学报，2000,35(8)：561～565．
[41]熊杰；冯亦璞．丁基苯酞对局灶性脑缺血过程中线粒体损伤的保护作用．药学学报，2000，
35(6)：408—412．
[42]董高翔；冯亦璞．丁基苯酞对局部脑缺血再灌注大鼠脑线粒体ATPase，抗氧化酶活性和脂
质过氧化的影响．中国医学科学院学报，2002,24(1)：93—97．
[43]种兆忠；冯亦璞．丁基苯酞对大鼠皮层神经元损伤后谷氨酸和5-羟色胺释放的影响．中国
药学杂志．1999,34(9)：589—591．
[44]胡盾；黄新祥；冯亦璞．丁基苯酞对全脑缺血大鼠的纹状体细胞外液嘌呤类代谢物含量的
影响．药学学报，1996,31(1)：13-17．
[45]种兆忠；冯亦璞丁基苯酞对大鼠局灶性脑缺血和重灌后脑内TXB-2和6-keto-PGF_(1a)含量
的影响中国药理学报1997，8(6)：505-508．
[46]Peng SH, Zhou TH. The study on in vitro metabolism of butylphthalide by gas chromatography-mass spectrometry. Chin Chem Lett, 1995, 6:55.
[47]彭仕华；周同惠．丁基苯酞的体内代谢转化研究．药学学报，1996,31(10)：780-784．
[48]王春华；冯亦璞；吴元鎏．丁基苯酞在大鼠中代谢产物的研究．药学学报，1997,32(9)：641-646．
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