HPLC法测定复方地龙胶囊中尿嘧啶、次黄嘌呤和肌苷的含量

目的：建立复方地龙胶囊中尿嘧啶、次黄嘌呤和肌苷的含量测定方法。方法：采用HPLC法,色谱柱以十八烷基硅烷键合硅胶为填充剂（250mm×4.6mm,5μm）,甲醇与含0.10%磷酸的0.050mol·L-1的磷酸二氢钾溶液为流动相梯度洗脱,检测波长254nm,流速1mL·min-1,柱温15℃。结果：尿嘧啶1.34～40.5ng范围内（r=1.0000）有良好的线性关系;次黄嘌呤在2.75～82.5ng范围内（r=1.0000）有良好的线性关系;肌苷在5.10～153.0ng范围内（r=1.0000）有良好的线性关系。平均回收率分别为97.96%、98.31%和97.96%;RSD分别为1.23%、0.78%和0.83%（n=6）。结论：该方法操作简单,为全面控制复方地龙胶囊的质量提供了可行性。     

喷施化肥和有机肥对山药成分的影响研究＊

目的 比较并评价喷施化肥和有机肥后对山药成分的影响。方法 分别对河北安国种植的小白嘴山药分3次喷施化肥尿素和有机肥，然后收集样品，采用烘干法测定其中水分的含量，通过凯氏定氮法检测蛋白质的含量，自动氨基酸分析仪测定氨基酸的含量，苯酚-硫酸比色法检测多糖的含量，高效液相色谱二极管阵列检测器联用仪（high performance liquid chromatography-diode array detector，HPLC-DAD）测定尿囊素、尿嘧啶和腺苷的含量。结果 施加化肥的山药中水分、蛋白质、尿嘧啶、腺苷和尿囊素的含量较高，而施加有机肥的山药中多糖含量较高，存在一定差异性。结论 对山药种植过程中施加不同肥料，可以影响其成分含量，进而影响其质量。     

HPLC法测定不同产地地龙中尿嘧啶、次黄嘌呤、尿苷、肌苷的含量

目的:测定不同产地、品种的地龙药材中尿嘧啶、次黄嘌呤、尿苷、肌苷的含量,建立地龙质量控制的客观指标。方法:用0.9%生理盐水超声提取地龙药材,采用反相高效液相色谱法,Waters Symmetry C₁₈色谱柱（150mm×4.6mm,5μm）,以0.01mol·L^-1磷酸二氢钾溶液和50%甲醇为流动相,流速为0.8ml·min^-1,梯度洗脱,检测波长254nm,柱温27℃,同时测定尿嘧啶、次黄嘌呤、尿苷、肌苷等4种成分的含量。结果:尿嘧啶的线性范围为0.75～24.00μg·ml^-1（r=0.9993）,平均回收率99.82%,RSD=2.34%（n=6）;次黄嘌呤的线性范围2.50～80.00μg·ml^-1（r=0.9992）,平均回收率101.93%,RSD=1.45%（n=6）;尿苷的线性范围1.25～40.00μg·ml^-1（r=0.9992）,平均回收率97.53%,RSD=1.73%（n=6）;肌苷的线性范围5.00～160.00μg·ml^-1（r=0.9991）,平均回收率102.06%,RSD=2.44%（n=6）。结论:该方法重复性,回收率好,可用于地龙药材中尿嘧啶等4种成分的含量测定。     

Human cytomegalovirus infection modulates DNA base excision repair in fibroblast cells

Regulation of DNA repair mechanisms during the viral replication cycle may have consequences for the virus with regards to genomic variability, adaptation, and replication of viral DNA. We have studied the activities and expression patterns of key enzymes involved in the first two steps of base excision repair (BER) of DNA in primary fibroblasts infected by human cytomegalovirus (HCMV). Infected cells were very proficient for removal of uracil and 5' hydrolysis of AP sites (AP endonuclease activity) as compared to the mock-infected cells, suggesting a direct role in generating free ends at uracil lesions in DNA for initiation of viral replication. Furthermore, the capacity to initiate repair of alkylated and oxidized base lesions were reduced in HCMV-infected cells, indicating increased mutation frequencies that could promote genetic variability. We hypothesize that modulation of BER activities may play an important role in HCMV pathogenesis to ensure efficient replication and genomic variation of viral DNA.     

Constituents of the Halophyte Salicornia herbacea

Four compounds were isolated from Salicornia herbacea by repeated column chromatography. Their structures were identified as beta-sitosterol (1), stigmasterol (2), uracil (3), and isorhamnetin-3-O-beta-D-glucopyranoside (4) by spectral analysis and comparison with the published data.     

基因扩增技术中抗污染的研究

目的　了解残留UDG对dU -DNAPCR产物的影响及灭活UDG时对Taq -P的影响。方法　采用微孔板杂交技术检测dU -DNAPCR产物 3 7℃放置 2 0h杂交百分率 ;检测在 -2 0℃、4℃、室温、3 7℃时不同保存条件下的杂交百分率 ;94℃灭活 10min对Taq -P活性的影响。结果　加 0 2单位、1 0单位UDG组比不加UDG组杂交A值下降 13 2 81%～ 2 0 5 5 7%。t检验差异有显著性 (t=2 85 5 ,2 869,P <0 0 5 ) ;经 94℃灭活产物组 3 7℃杂交A值比 -2 0℃A值下降 5 5 47% ,未经 94℃灭活杂交A值下降 10 3 45 % ;94℃预变性 3 0s杂交A值为 98 714 %、10min 3 0s组杂交A值为 96 818%。结论　以上结果提示经 94℃加热灭活对PCR产物保存有重要作用 ,不仅对灭活UDG有作用而且还可灭活来源于UDG以外可破坏DNA的酶。表明含UDG组杂交A值比不加UDG组低13 2 81%～ 2 0 5 5 7%与UDG有关。     

mdr1启动子调控双自杀基因对卵巢癌SKOV3耐药细胞的靶向杀伤作用

目的观察多药耐药基因1(mdr1)启动子调控腺病毒介导的胞嘧啶脱氨酶∷尿嘧啶磷酸核糖转移酶(CD∷UPP)融合基因系统对人卵巢癌紫杉醇耐药细胞的靶向杀伤作用。方法2004年10月至2005年5月于华中科技大学附属协和医院,将重组腺病毒(Admdr1-CD∷UPP)转染卵巢癌紫杉醇耐药细胞(SKOV3/Taxol)及非耐药细胞(SKOV3),荧光显微镜下观察转染效率,用逆转录-聚合酶链反应(RT-PCR)检测目的基因CD∷UPP的表达,并给予不同浓度的5-氟胞嘧啶(5-FC),用四甲基偶氮唑蓝(MTT)法检测二组的细胞存活率及旁观者效应。结果重组腺病毒对SKOV3/Taxol的转染率随腺病毒滴度的递增而增加,感染复数(MOI)为50时,感染率约100%;CD∷UPP基因仅在SKOV3/Taxol中稳定表达;5-FC对耐药细胞转基因组的杀伤作用显著高于非耐药细胞转基因组,前者表现出明显的旁观者效应。结论mdr1启动子调控的CD∷UPP融合自杀基因系统对人卵巢癌紫杉醇耐药细胞有靶向杀伤作用。     

Modification by Analgesics of Lesion Development in the Urinary Tract and Various Other Organs of Rats Pretreated with Dihydroxy-di-N-propylnitrosamine and Uracil

Effects of the analgesics phenacetin, acetaminophen and antipyrine on lesion development in the urinary tract and other organs in male F344 rats were investigated. Animals were concurrently administered with 0.1% dihydroxy-di-N-propylnitrosamine (DHPN) in drinking water and 3.0% uracil in the diet for 4 weeks and then, starting 1 week after the cessation of this treatment, received basal diet or diet containing phenacetin, acetaminophen or antipyrine for 35 weeks. The occurrences of renal cell tumors were increased in the groups given phenacetin or antipyrine, as compared with the DHPN+uracil alone controls. Antipyrine, but not the two other compounds, also enhanced development of hyperplastic lesions in the renal pelvis and ureter. In the urinary bladder, phenacetin and antipyrine treatments were both associated with increased incidence of preneoplastic or neoplastic lesions. Furthermore, phenacetin alone, without the initiating agent pretreatments, induced simple hyperplasias of the urinary bladder at high incidence. Antipyrine enhanced induction of hyperplastic lesions in the ureter and was also found to increase the incidences of preneoplastic and neoplastic lesions in the liver. Although decreased incidences of tumor development of lung and thyroid were observed for the group given phenacetin, this might have been linked to the decreased weight gain. The results confirmed that combination treatment with DHPN+uracil is effective for wide-spectrum initiation of carcinogenesis in the urological tract and demonstrated significant modification potential for both phenacetin and antipyrine.     

胞嘧啶脱氨酶尿嘧啶-磷酸核糖基转移酶融合基因/5-氟胞嘧啶治疗鼻咽癌的实验研究

目的研究胞嘧啶脱氨酶-尿嘧啶磷酸核糖基转移酶融合基因/5-氟胞嘧啶(fusion cytosine deaminase- uracil phosphoribosyl transferase gene/5-Flurocy- tosine,FCU/5-FC)对鼻咽癌CNE-2细胞的杀伤效应以及对鼻咽癌移植瘤生长的抑制作用。方法构建表达质粒载体pcDNA3．1(-)CMVe·Egr-1·FCU,BamHⅠ酶切鉴定重组质粒,分析FCU基因和融合启动子CMVe·Egr-1的序列。电穿孔法将重组质粒转染CNE-2细胞,300μg/ml～600μg/mlG418筛选14天获得稳定表达FCU基因的CNE-2细胞,RT-PCR鉴定FCU基因的表达。以转染空白载体pcDNA3．1(-)的CNE-2细胞为对照,MTT法观察5-FC对表达FCU基因的CNE-2细胞生长的抑制作用,考察FCU基因的细胞旁杀效应。5×10~6的CNE-2细胞接种裸鼠右腋下,建立鼻咽癌裸鼠移植瘤模型,12天后将裸鼠随机分成3组：CU组,CU+PBS组和CU+5-FC组,观察不同处理对移植瘤生长的抑制作用。结果酶切和序列分析证明重组质粒含完整的FCU基因和Egr-1启动子的核心序列,RT-PCR从转染细胞总RNA中扩出330bp的预期片段。表达FCU基因的CNE-2细胞在5-FC干预下,生长受到抑制。随着表达FCU基因的细胞比例增加,细胞存活率相应地下降。CU组,CU+PBS组移植瘤体积与CU+5-FC组有显著性差异(P＜0．05)。结论表达FCU基因的CNE-2细胞可以被5-FC杀伤,FCU/5-FC系统抑制了移植瘤生长,为治疗鼻咽癌提供了新的策略。     

Adsorption of uracil on bismuth single crystal planes

Uracil adsorption at Bi(h k l) has been studied using impedance spectroscopy method. The limiting adsorption potential shift, Gibbs energy of adsorption $-\mathrm{\Delta }{G}_{\mathrm{A}}^{°}$ and interaction constant values a in the Frumkin isotherm have been calculated and compared with the corresponding data for Hg [V. Brabec, S.D. Christian, G. Dryhust, J. Electroanal. Chem. 85 (1977) 389; V. Brabec, S.D. Christian, G. Dryhust, Biophys. Chem. 7 (1978) 253]. It was found that the values of $|-\mathrm{\Delta }{G}_{\mathrm{A}}^{°}|$ increase in the order $\mathrm{Hg}<\mathrm{Bi}(111)<\mathrm{Bi}(01\overline{1})$, but the values of a are higher for Bi(h k l) compared with Hg [V. Brabec, S.D. Christian, G. Dryhust, J. Electroanal. Chem. 85 (1977) 389; V. Brabec, S.D. Christian, G. Dryhust, Biophys. Chem. 7 (1978) 253]. The values of the limiting adsorption potential shift are comparable for the Hg and Bi(h k l) electrodes.