^1H核磁共振谱直接观测N,N—二甲氨基甲酸—间—（2—二甲…

应用高分辨＾１Ｈ核磁共振谱直接观测Ｎ，Ｎ－二甲氨基甲酸－间－（２－二甲氨基）乙氧基苯酯在原位灌流大鼠肝脏中的代谢产物和代谢动力学过程。灌流不同时间采集样本在４００ＭＨＺＮＭＲ谱仪上直接用选择自旋翻转回波＾１ＨＮＭＲ法进行定性和定量分析。ＤＭＤＭＣ在灌流大鼠肝肘中的主要代谢产物是羟基ＤＭＤＭＣ和Ｎ－去甲基ＤＭＤＭＣ，其消除半衰期为１１２ｍｉｎ。     

可逆性胆碱酯酶抑制剂 Ⅱ.二甲氨基甲酸间-(烷氨基)烷硫基苯酯的合成

前文报道了一系列二甲氨基甲酸间-(烷氨基)烷氧基苯酯(I,X=O)的合成,发现催醒安(I,X=O,n=2,R=CH_3)对临床中麻催醒有较好的效果,是一个结构简单、易于合成、具有中枢作用的可逆性胆碱酯酶抑制剂。但催醒安对中枢胆碱酯酶的抑制作用不够强,为此,我们合成了相应的硫代衍生物(Ⅱ,X=S)(表2)以期能增强对中枢胆碱酯酶的抑制作用。     

可逆性胆碱酯酶抑制剂:二甲氨基甲酸-[2-(2-二甲氨基)乙氧基-4(或5)-特丁基]苯酯的合成

前报报道在“催醒安”的对位异构体的苯环上引入烷基,可以增强对胆碱酯酶的抑制作用。为了探索在邻位异构体(Ⅰ)的苯环上引入烷基对生理活性的影响,合成了二甲氨基甲酸-[2-(2-二甲氨基)乙氧基-4(或5)-特丁基]苯酯(Ⅱ)。     

可逆性胆碱酯酶抑制剂:二甲氨基甲酸-[3-烷基-4-(烷氨基)烷氧基]苯酯的合成

前文报道的一系列二甲氨基甲酸间-(烷氨基)烷氧(烷硫)基苯酯类化合物(I,X=O,S)中,多数具有较强的抑酶作用,有的对实验动物的中麻催醒效果较好,且毒性和副作用小。其中二甲氨基甲酸间-(2-二甲氨基)-乙氧基苯酯(I,X=O,n=2,R=CH_3)已命名为“催醒     

可逆性胆碱酯酶抑制剂二甲氨基甲酸-[3-(烷氨基)烷氧基-4(5)-叔丁基]苯酯的合成

在催醒安的邻位和对位异构体及其同系物的苯坏上引入烷基后,可以增强抑制胆碱酯酶的活性,我们乃进一步在催醒安及其同系物的苯环不同位置上引入叔丁基,合成了一系列二甲氨基甲酸-[3-(烷氨基)烷氧基-4(5)-叔丁基]苯酯(Ⅰ_(1～13)和Ⅱ_(1～5))(表2),以探索对活性的影响。     

N,N-二甲氨基甲酸5-(1,3,3-三甲基吲哚满)酯及其代谢产物在大鼠离体肝脏中的代谢动力学

利用大鼠肝脏灌流技术研究了N,N-二甲氨基甲酸5-(1,3,3-三甲基吲哚满)酯(TMDMC)及其代谢产物在大鼠离体肝脏中的代谢动力学,TMDMC在改良的灌流液中作循环式灌流,于不同时间留取少量灌流液,用高效液相色谱(HPLC)作药物的定量分析,结果显示其在离体灌流肝脏中的清除模型符合两相消除模型,动力学参数表明,TMDMC在肝内的分布较快,其分布半衰期仅为4min,肝脏分布容积为102.4 ml,清除率是0.67ml·min~(-1),清除比率为0.027.灌流2h时,其浓度已下降了73％.与此同时,TMDMC的代谢产物逐渐生成,其中以产物Ⅲ的生成量最大,其最高生成浓度(C_(max))为447.1 μg·ml~(-1),2 h产物Ⅲ的生成总量已达TMDMC灌流量的25.6％.而代谢产物Ⅱ,Ⅵ的生成较少,其C_(max)分别是20.4μg·ml~(-1)和20.7μg·ml~(-1).     

N－（4－甲氧羰基－4－邻苯二甲酰亚氨基丁酰基）－N－取代甘氨酸、脯氨酸和焦谷氨酸的合成

报道ｌｌ个预期有血管紧张素转化酶抑制活性的Ｎ－（４－甲氧羰基－４－邻苯二甲酰亚氨基丁酰基）－Ａ－取代甘氨酸（ＶＩＩ１－９）、脯氨酸（ＶＩｌ１０）和焦谷氨酸（ＶＩｌ１１）的合成和鉴定。所有上述化合物以及与ＶＩＩ１－９相应的叔丁酯（ＶＩ１～９）均未见文献报道。药理初试结果显示，化合物ＶＩＩ８，ＶＩＩ９和ＶＩｌ１０均有明显降压活性。     

可逆性胆碱酯酶抑制剂:二甲氨基甲酸-5-(1,3,3-三甲基-6-取代基)吲哚满酯的合成

催醒宁(I,R=H,R′X=HCl)对胆碱酯酶的抑制作用较强,但稳定性较差,作用时间较短。推测可能与其酯基易于水解有关。鉴于催醒宁结构中的氨基甲酸酯是抑酶作用的药效基团,酯基被水解后,抑酶活性即消失,我们设想,如果在催醒宁结构中的酯基邻位上引入取代基,利用其空间效应的影响,使酯基增加对水解的稳定性,或有可能达到延长作用时间的目的。因此,我们合成了二甲氨基甲酸-5-(1,3,3-三甲基-6-取代基)吲哚满酯盐酸盐和季铵盐(Ⅰ_(1～14),表1),以探索取代基对抑酶强度和作用时间的影响。     

抗肿瘤药物2，3－二乙酰氧基－5－烷氧羰基－1－（3′，5′－二酮－N（4′）－取代哌嗪甲基）苯的合成研究（Ⅲ）

在寻找新型抗肿瘤药物过程中，作者设计并合成了十四个３．５－哌嗪二酮类化合物。经过对肿瘤细胞Ｐ３８８体外试验结果表明，化合物（Ⅲ１，Ⅲ５，Ⅲ７）活性较高，其他化合物也只有一定的生物活性．（Ⅲ１：剂量：１γ／ｍＬ，抑制率：５０％：Ⅲ５：剂量：１γ／ｍＬ，抑制率：７３％；Ⅲ７：剂量：１γ／ｍＬ，抑制率：７２％）     

~(19)F体内核磁共振技术观测5-氟尿嘧啶在小鼠肝脏中的代谢

应用１９Ｆ体内核磁共振技术（１９ＦＮＭＲ）直接观测了５－氟尿嘧啶（５－ＦＵ）在小鼠肝脏中的代谢产物和代谢动力学，昆明种（ＫＭ）小鼠和Ｃ５７小鼠ｉｖ５－ＦＵ２００ｍｇ·ｋｇ－１后，用１９Ｆ表面线圈测得５－ＦＵ在肝脏中依次代谢为α－氟－β－脲基丙酸（ＦＵＰＡ）和α－氟－β－丙氨酸（ＦＢＡＬ）．在荷Ｓ１８０瘤ＫＭ小鼠肝脏中还可检测到活化产物氟代核苷／核苷酸（ＦＮＵＣ）．５－ＦＵ在ＫＭ和Ｃ５７小鼠肝脏中清除较快，消除半衰期分别为１６．７±３．０和３６．６±２．４ｍｉｎ，其主要产物ＦＢＡＬ的最大生成ｒｍａｘ（相对强度比）分别为８４±１４和９２±１０．但在ＫＭ小鼠肝脏中由ＦＵＰＡ进一步降解为ＦＢＡＬ的速率明显快于Ｃ５７小鼠，ｔ１／２ｒ分别为３１±８和５７±１３ｍｉｎ．相应地，在Ｃ５７小鼠肝脏中ＦＵＰＡ能维持一定的水平并达到ｒｍａｘ２６．２±２．２．     

19F体内核磁共振技术观测5-氟尿嘧啶在小鼠肝脏中的代谢

应用¹⁹F体内核磁共振技术(¹⁹F NMR)直接观测了5-氟尿嘧啶(5-FU)在小鼠肝脏中的代谢产物和代谢动力学，昆明种(KM)小鼠和C57小鼠iv 5-FU 200 mg·kg^-1后，用¹⁹F表面线圈测得5- FU在肝脏中依次代谢为α-氟-β-脲基丙酸(FUPA)和α-氟-β-丙氨酸(FBAL). 在荷S180瘤KM小鼠肝脏中还可检测到活化产物氟代核苷/核苷酸(FNUC). 5-FU在KM和C57小鼠肝脏中清除较快，消除半衰期分别为16.7±3.0和36.6±2.4 min，其主要产物FBAL的最大生成r_max(相对强度比)分别为84±14和92±10. 但在KM小鼠肝脏中由FUPA进一步降解为FBAL的速率明显快于C57小鼠，t_1/2r分别为31±8和57±13 min. 相应地，在C57小鼠肝脏中FUPA能维持一定的水平并达到r_max 26.2±2.2.     

2-苯甲酰肼叉-1,3-二噻茂烷在大鼠原位灌流肝中的代谢动力学

采用反相高压液相色谱的三内标三波长切换技术对不同剂量的农药2-苯甲酰肼叉-1.3二噻茂烷在大鼠原位灌流肝中的代谢动力学进行了研究.结果表明,该农药经门静脉进入大鼠原位灌流肝后,很快分布于肝脏中,而在灌流肝中的消除过程较缓慢。随着给药剂量的增加,大鼠肝灌流液中各种代谢产物的生成量也逐渐增加,尤以肼叉1.3-二噻茂烷和苯甲酸生成量的增加更为显著.可见,该农药在大鼠原位灌流肝中的主要代谢途径为水解作用。     

Metabolism of 14C-2',3'-dideoxyinosine by the in situ perfused rat liver preparation

The metabolism and biliary excretion of 14C-dideoxyinosine (14C-ddI) has been investigated using the in situ perfused rat liver (PRL) preparation. After 2 h of perfusion through the liver, approximately 70-75 per cent of the total 14C-radiolabel was recovered in the perfusion medium, less than 1 per cent was excreted in bile and 15-18 per cent was retained in the liver. Hepatic clearance of ddI was 1.5 +/- 0.1 ml min-1 and half-life for the elimination of ddI from the medium was 22.9 +/- 2.0 min (n = 3). Hepatic extraction was estimated to be 7.5 per cent. HPLC analysis of the effluent perfusate indicated that ddI was metabolized to hypoxanthine, xanthine, uric acid, and to a polar metabolite which was tentatively identified as allantoin. Approximately 60-65 per cent of the ddI dose was converted to allantoin after 2 h of perfusion. Of the other three metabolites, uric acid levels increased to 20-30 per cent of the dose after 45 min and declined to about 5 per cent of the dose by the end of the perfusion period. Levels of hypoxanthine and xanthine were low and both compounds were not detected in the perfusate after 45 min post-infusion. In bile, the major peak, which accounted for about 50 per cent of the 14C-radiolabel co-eluted with the putative metabolite, allantoin (0.4 per cent of the dose). Uric acid (0.06 per cent of the dose) was the only other metabolite detected in bile. These results suggest that biliary excretion is a minor pathway for the elimination of ddI. Furthermore, ddI is rapidly cleared and metabolized by the liver to hypoxanthine, xanthine, uric acid, and to allantoin.     

The effect of three H2 receptor antagonists on the disposition of cyclosporin a in the in situ perfused rat liver model

The in situ, perfused rat liver model was used to investigate the effect of three H₂ receptor antagonists on the disposition of cyclosporin A (CyA) and the major human metabolite, AM1. Perfusion experiments, using standard techniques, were carried out on four groups (one control and three H₂-receptor antagonist-treated groups) of male Sprague-Dawley rats (300–350 g). All animals received CyA, 2.5 mg; the three treated groups received cimetidine (8 mg), ranitidine (3 mg), or famotidine (0.4 mg). Perfusated and bile samples were collected and assayed for CyA, AM1, and the H₂ receptor antagonists by HPLC. Results indicated that CyA perfusate concentrations in the controls and cimetidine and ranitidine-treated groups were not significantly different, although levels in the famotidine group were significantly higher at all times (p<0.05), except 30 min, compared to the controls. However, examination of the AM1 perfusate and bile data and the apparent metabolic clearance data indicated that CyA metabolism was still occurring, despite the presence of the H₂ receptor antagonist. It is suggested that the absence of a interaction may be attributed to a lack of specificity of the H₂ receptor antagonists for CYP3A, the isoenzyme responsible for CyA metabolism.     

Effects of Di-2-ethylhexyl phthalate (DEHP) on gap-junctional intercellular communication (GJIC), DNA synthesis, and peroxisomal beta oxidation (PBOX) in rat, mouse, and hamster liver.

The present study evaluated the effect of di-2-ethylhexyl phthalate (DEHP) on gap-junctional intercellular communication (GJIC), peroxisomal beta-oxidation (PBOX) activity, and replicative DNA synthesis in several rodent species with differing susceptibilities to peroxisome proliferator-induced hepatic tumorigenesis. A low (non-tumorigenic) and high (tumorigenic) dietary concentration of DEHP was administered to male F344 rats for 1, 2, 4, and 6 weeks. Additionally, a previously non-tumorigenic dose (1000 ppm) and tumorigenic dose of DEHP (12,000 ppm), as determined by chronic bioassay data, were examined following 2 weeks dietary administration. Male B6C3F1 mice were fed the non-tumorigenic concentration, 500 ppm, and the tumorigenic concentration, 6000 ppm, of DEHP for two and four weeks. The hepatic effects of low and high concentrations of DEHP, 1000 and 6000 ppm, were also examined in male Syrian Golden hamsters (refractory to peroxisome proliferator-induced tumorigenicity). In rat and mouse liver, a concentration-dependent increase in the relative liver weight, PBOX activity, and replicative DNA synthesis was observed at the earliest time point examined. Concurrent to these observations was an inhibition of GJIC. In hamster liver, a slight increase in the relative liver weight, PBOX activity, and replicative DNA synthesis was observed. However, these effects were not of the same magnitude or consistency as those observed in rats or mice. Furthermore, DEHP had no effect on GJIC in hamster liver at any of the time points examined (2 and 4 weeks). HPLC analysis of DEHP and its primary metabolites, mono-2-ethylhexyl phthalate (MEHP), and phthalate acid (PA), indicated a time- and concentration-dependent increase in the hepatic concentration of MEHP. At equivalent dietary concentrations and time points, the presence of MEHP, the primary metabolite responsible for the hepatic effects of DEHP, demonstrated a species-specific response. The largest increase in the hepatic concentration of MEHP was observed in mice, which was greater than the concentration observed in rats. The hepatic concentration of MEHP was lowest in hamsters. Hepatic concentrations of DEHP and phthalic acid were minimal and did not correlate with concentration and time. Collectively, these data demonstrate the inhibition of hepatic GJIC and increased replicative DNA synthesis correlated with the observed dose- and species-specific tumorigenicity of DEHP and may be predictive indicators of the nongenotoxic carcinogenic potential of phthalate esters.     

Labelling of I2B-imidazoline receptors by [3H]2-(2-benzofuranyl)-2-imidazoline (2-BFI) in rat brain and liver: characterization, regulation and relation to monoamine oxidase enzymes

The novel selective imidazoline radioligand [3H]2-(2-benzofuranyl)-2-imidazoline (2-BFI) was used to characterize and assess further the nature of I₂-imidazoline receptors in rat brain and liver. In the cerebral cortex, 2-BFI displayed high affinity (K _i= 9.8 nM) for a single class of [³H]2-BFI binding sites. Other imidazoline/guanidine compounds (e.g. aganodine, cirazoline and idazoxan) displayed biphasic competition curves, indicating the existence of high (K _iH= 2.9-78 nM; R _H= 61-83%) and low (K _iL= 4.7-158 μM) affinity sites. The pharmacological profile for [³H]2-BFI binding (aganodine > cirazoline > 2-BFI >> clonidine > amiloride >> efaroxan) was typical of that for I₂-sites. This profile was almost identical to that obtained against [³H]idazoxan (correlation between pK _i values, r = 0.97) which indicated that the sites characterized with [³H]2-BFI in brain corresponded to I₂-imidazoline receptors. The low affinity of amiloride against [³H]2-BFI (K _i= 900 nM) further indicated that these brain I₂-sites belong to the I_2B-subtype. [³H]2-BFI binding sites (B _max= 72 fmol/mg protein) in brain were differentially modulated by treatment (7 days) with cirazoline (up-regulation: 25%) and the MAO inhibitor phenelzine (down-regulation: 31%), indicating that these I₂-sites are regulated in vivo, as is the case for those labelled by [³H]idazoxan. Chronic treatment with 2-phenylethylamine, a phenelzine metabolite and endogenous amine, did not alter the density of brain of I₂-imidazoline receptors labelled by [³H]idazoxan. Preincubation of liver membranes with the MAO inhibitor clorgyline (10^-7 M) abolished the binding of [³H]Ro 41-1049 (N-(2-aminoethyl)-5-(m-fluorophenyl)-4-thiazole carboxamide) to MAO-A, but it did not alter the binding of [³H]Ro 19-6327 (N-(2-aminoethyl)-5-chloro-2-pyridine carboxamide) to MAO-B or that of [³H]2-BFI to I₂-sites. At 10^-4 M it also abolished MAO-B sites, but a substantial proportion of I₂-sites (40%) remained intact. Preincubation of liver membranes at 60 °C also abolished MAO-A/B sites, whereas still 22% of I₂-sites remained. The results indicate that [³H]2-BFI is a good tool for the identification of I₂-imidazoline receptors and suggest further that certain I₂-sites and MAO are different proteins. Received: 10 January 1997 / Accepted: 6 March 1997     

Localization of somatostatin (SRIF) SSTR-1, SSTR-2 and SSTR-3 receptor mRNA in rat brain by in situ hybridization

In situ hybridization histochemistry was performed to analyse the distribution of the messenger RNA (mRNA) of three putative somatostatin (SRIF) receptors in rat brain, using oligonucleotide probes derived from the cDNA coding for SSTR-1, SSTR-2, and SSTR-3 receptors.SSTR-1 signals were found in layers VVI of the cerebral cortex, in primary olfactory cortex, taenia tecta, subiculum, entorhinal cortex, granular layer of the dentate gyrus, amygdala and cerebellar nuclei. Signals for SSTR-2 were found in the frontal cerebral cortex (layers IV, V and VI), taenia tecta, claustrum, endopiriform nucleus, locus coeruleus, medial habenula, subiculum, granular cell layer of the dentate gyrus and amygdala. High levels of SSTR-3 hybridization were found in the olfactory bulb, primary olfactory cortex, islands of Calleja, medial habenula, amygdala, granular layer of the dentate gyrus, various thalamic and pontine nuclei and in the granular and Purkinje cell layers of the cerebellum.The distribution of the hybridization signals of the oligoprobes is consistent with the labelling of specific SRIF binding sites in rat brain. Especially, SSTR-2 and SSTR-1 oligos seem to label regions in which SS-1 and SS-2 receptors, respectively, have been previously characterized in autoradiographical studies. The situation is less clear with SSTR-3 mRNA, since SRIF binding in adult rats is usually low or absent in cerebellum, although some cerebellar nuclei appear to be labelled in the adult. The localization of SSTR-1, SSTR-2 and SSTR-3 mRNAs suggests that SRIF receptor subtypes in rat brain show profound differences in their distribution and are involved in a variety of central, in addition to neuroendocrine, functions.Monique Rigo, who contributed very significantly to this work, died tragically on January 21, 1993 Correspondence to: D. Hoyer at the above address     

Deacetylation of 4-(5-acetylamino-2-furyl)thiazole and formation of 1-(4-thiazolyl)-3-cyano-1-propanone by rat liver tissues

The reductive metabolism of the rat carcinogen 4-(5-nitro-2furyl)thiazole (NFT) to 1-4-thiazolyl)-3-cyano-1-propanone (TCP) is reported. Formation of TCP from NFT involved furan ring fission. This could have occurred through involvement of either aminofuran or N-hydroxylaminofuran as precursors. To examine if 4-(5-amino-2-furyl)thiazole is a precursor for TCP, a stable model compound, 4-(5-acetylamino-2-furyl)thiazole (AAFT), was prepared and subjected to enzymatic deacetylation, using rat liver tissue homogenates. AAFT was synthesized by catalytic hydrogenation of NFT with 5% palladium on activated carbon, followed by acetylation with acetic anhydride. AAFT, a white crystalline powder, melted at 168–170°, had an extinction coefficient of 17.9 mM^?1 cm^?1 at 293 nm in ethyl acetate, and exhibited spectroscopic and mass spectral characteristics consistent with the assigned structure. Incubation with rat liver 10,000 g supernatant preparations resulted in the biotransformation of AAFT as evidenced by a decrease in absorption at 290 nm. Incubation of ¹⁴C-labeled AAFT followed by extraction with chloroform-diethyl ether (1:1) resulted in the recovery of a major portion (56%) of the radioactivity in the organic phase when the label was at the 2-position of the thiazole ring, while the major amount (82%) of radioactivity was recovered in the aqueous phase when the 1-¹⁴C-acetyl group was labeled. The radioactivity from the aqueous phase was extractable into the organic phase following acidification to pH 1, an observation consistent with deacetylation. Furthermore, the deacetylation product exhibited a mass spectrum, and retention times in gas and high pressure liquid chromatography, similar to those of synthetic TCP. These data establish 4-(5-amino-2-furyl)thiazole, derived from AAFT by deacetylation, as a precursor for TCP.