顶空气相色谱法测定注射用胸腺法新中5种有机溶剂残留量

目的 建立同时测定注射用胸腺法新中乙腈、二氯甲烷、醋酸乙酯、苯和苯甲醚5种有机溶剂残留量的顶空气相色谱法。方法 采用Agilent DB-624（30 m×0.53 mm×3 μm）毛细管色谱柱;火焰离子化检测器;进样口温度200℃;检测器温度250℃;载气为氮气;载气体积流量为2.0 mL·min^-1;分流比为10：1;升温程序：起始温度40℃,保持6 min,以8℃·min^-1的速率升温至90℃,保持2 min,再以20℃·min^-1的速率升温至200℃,保持5 min;采用顶空进样方式,顶空加热箱温度80℃,样品瓶平衡时间30 min。进行系统适用性、检测限（LOD）与定量限（LOQ）、线性关系和范围、加样回收率、精密度、稳定性、耐用性考察。结果 注射用胸腺法新中5种残留溶剂在各自线性浓度范围内与峰面积线性关系良好;平均加样回收率在95.7%~106.3%;精密度、稳定性、耐用性均符合要求。5批胸腺法新中均未检出5种有机溶剂。结论 建立的顶空气相色谱法操作简单、灵敏度和准确度高、重现性和耐用性好,可用于注射用胸腺法新中5种有机溶剂残留量的测定。     

Analgesic, anti-inflammatory and antioxidant properties of Buddleja globosa, Buddlejaceae

ETHNOPHARMACOLOGICAL RELEVANCE: Buddleja globosa, known as "matico", is employed in Chile for wound healing. AIM OF THE STUDY: To validate the traditional use of the crude drug through in vivo and in vitro evaluation of the anti-inflammatory, analgesic and antioxidant properties of its extracts. MATERIALS AND METHODS: Sequential hexane, dichloromethane, methanol and total methanol extracts were studied using bioguided fractionation. The following activities were investigated: analgesic (writhing test), oral and topic anti-inflammatory (paw- and ear-induced edema), free radical scavenging and antioxidant activities (1,1-diphenyl-2-picrylhydrazyl, DPPH, superoxide anion, lipid peroxidation and xanthine oxidase inhibition). Sodium naproxen, nimesulide, indomethacin were used as reference drugs for in vivo, quercetin and allopurinol for in vitro assays. RESULTS: A mixture of alpha- and beta-amyrins was isolated from the hexane extract that showed 41.2% of analgesic effect at 600 mg/kg, inhibited by 47.7 and 79.0% the arachidonic acid (AA) and 12-deoxyphorbol-13-decanoate (TPA)-induced inflammation at 3mg/20 microL/ear, respectively. A mixture of beta-sitosterol, stigmasterol, stigmastenol, stigmastanol and campesterol was isolated from the fraction CD4-N and beta-sitosterol-glycoside from the fraction CD5-N, reducing TPA-induced inflammation by 78.2 and 83.7% at 1mg/20 microL/ear, respectively. The fraction CD4-N at 300 mg/kg also showed analgesic activity (38.7%). The methanol extract at 600mg/kg per os showed anti-inflammatory effect (61.4%), topic anti-inflammatory (56.7% on TPA) and analgesic activity (38.5%). Verbascoside and luteolin-7-O-glucoside were the major components of the methanol extract; apigenin 7-O-glucoside was also detected. Inhibition of superoxide anion, lipoperoxidation, and DPPH bleaching effect was found in the methanol serial and global extracts. CONCLUSIONS: The present report demonstrate the analgesic and anti-inflammatory properties of Buddleja globosa and validate its use in Chilean traditional medicine.     

Nobiletin and its derivatives overcome multidrug resistance (MDR) in cancer: total synthesis and discovery of potent MDR reversal agents

Our recent studies demonstrated that the natural product nobiletin (NOB) served as a promising multidrug resistance (MDR) reversal agent and improved the effectiveness of cancer chemotherapy in vitro. However, low aqueous solubility and difficulty in total synthesis limited its application as a therapeutic agent. To tackle these challenges, NOB was synthesized in a high yield by a concise route of six steps and fourteen derivatives were synthesized with remarkable solubility and efficacy. All the compounds showed improved sensitivity to paclitaxel (PTX) in P-glycoprotein (P-gp) overexpressing MDR cancer cells. Among them, compound 29d exhibited water solubility 280-fold higher than NOB. A drug-resistance A549/T xenograft model showed that 29d, at a dose of 50 mg/kg co-administered with PTX (15 mg/kg), inhibited tumor growth more effective than NOB and remarkably increased PTX concentration in the tumors via P-gp inhibition. Moreover, Western blot experiments revealed that 29d inhibited expression of NRF2, phosphorylated ERK and AKT in MDR cancer cells, thus implying 29d of multiple mechanisms to reverse MDR in lung cancer.     

Penta-block copolymer microspheres: Impact of polymer characteristics and process parameters on protein release

Here, we aimed to develop protein loaded microspheres (MSs) using penta-block PLGA-based copolymers to obtain sustained and complete protein release. We varied MS morphology and studied the control of protein release. Lysozyme was used as a model protein and MSs were prepared using the solid-in-oil-in-water emulsion solvent extraction method. We synthesized and studied various penta-block PLGA-based copolymers. Copolymer characteristics (LA/GA ratio and molecular weight of PLGA blocks) influenced MS morphology. MS porosity was influenced by process parameters (such as solvent type, polymer concentration, emulsifying speed), whereas the aqueous volume for extraction and stabilizer did not have a significant effect. MSs of the same size, but different morphologies, exhibited different protein release behavior, with porous structures being essential for the continuous and complete release of encapsulated protein. These findings suggest strategies to engineer the morphology of MSs produced from PLGA-based multi-block copolymers to achieve appropriate release rates for a protein delivery system.     

Intrahepatic cholangiocarcinoma in a worker at an offset color proof‐printing company: An autopsy case report

A 40‐year‐old Japanese man visited our hospital after test results indicated elevated hepatobiliary enzymes. He had worked at a printing plant for 8 years and been exposed to organic solvents, including 1,2‐dichloropropane (1,2‐DCP) and dichloromethane (DCM). Abdominal computed tomography (CT) showed an intrahepatic tumor with dilation of the intrahepatic bile duct. He was diagnosed with intrahepatic cholangiocarcinoma. He had no known risk factors for cholangiocarcinoma. Extended left hepatectomy with lymph node dissection was performed and the tumor was histologically diagnosed as well‐differentiated adenocarcinoma. A histological examination also showed biliary intraepithelial preneoplastic lesions in non‐cancerous liver areas. Two years after surgery, the patient developed jaundice, esophageal varices and ascites. A CT examination showed liver cirrhosis without recurrence of the cholangiocarcinoma. Although a liver transplantation was planned as a therapeutic option for his liver cirrhosis, his liver failure progressed rapidly and he died before transplantation could be performed. At autopsy, fibrosis was found in the whole liver, especially in the wall of the bile duct and periductal area suggesting chronic bile duct injury due to exposure to organic solvents. Taken together, the current case may suggest that exposure to organic solvents, including 1,2‐DCP and DCM, is a risk factor for cholangiocarcinoma. Identifying risk factors for cholangiocarcinoma will help identify the mechanism and help prevent development of the disease.     

吹扫捕集-气相色谱-质谱联用法测定生活饮用水中的二氯甲烷

目的建立生活饮用水中二氯甲烷的吹扫捕集-气相色谱-质谱联用测定方法。方法采用吹扫捕集富集水中二氯甲烷,解吸后用气相色谱-质谱测定,选择离子扫描方式下用标准曲线法进行定量分析。结果该方法操作简便,检出限低(0.005ng/mL),回收率均大于97%,相对标准偏差小于5%。结论该方法适合生活饮用水中的二氯甲烷的测定。     

Evaluation of concentration procedures of mutagenic metabolites from urine of diesel particulate-treated rats

Several concentration procedures of mutagenic metabolites contained in the urine of diesel particulate-treated rats were compared. Mutagenicity was monitored by the Salmonella/microsome assay. The procedures tested were: lyophilization; filtration on XAD-2, XAD-7 or Sephadex LH-20 matrices; ultrafiltration; and extraction with organic solvents. Urine extraction with dichloromethane (DCM) gave almost quantitative recovery of activity while leaving salts and other polar compounds in the aqueous phase, and is the method recommended.     

Lack of UDS activity in the livers of mice and rats exposed to dichloromethane

Dichloromethane (DCM) has been evaluated for its ability to initiate unscheduled DNA synthesis (UDS) in the livers of male mice and rats in vivo. Two types of experiment were conducted. In the first, Alpk:AP rats were exposed by oral gavage to 100, 500, or 1,000 mg/kg DCM and hepatocytes assessed for UDS via autoradiography 4 and 12 hours later. In the second, Fischer F344 rats or B6C3F1 mice were exposed by inhalation to either 2,000 or 4,000 ppm of DCM for either 2 or 6 hours, and hepatocytes assessed for UDS immediately after exposure. The dose levels and strains of rodent employed in the latter protocol correspond to those employed in a recent cancer bioassay of DCM conducted by the U.S. National Toxicology Program. DCM failed to induce UDS in any of the experiments. These data are discussed within the context of other evidence indicating DCM to be nongenotoxic in vivo, despite its reported carcinogenicity in the mouse.     

DNA adducts and oxidative DNA damage induced by organic extracts from PM2.5 in an acellular assay

The genotoxic activities of complex mixtures of organic extracts from the urban air particles collected in various localities of the Czech Republic, which differed in the extent and sources of air pollution, were compared. For this purpose, PM2.5 particles were collected by high volume samplers in the most polluted area of the Czech Republic - Ostrava region (localities Bartovice, Poruba and Karvina) and in the locality exhibiting a low level of air pollution - Trebon - a small town in the non-industrial region of Southern Bohemia. To prepare extractable organic matter (EOM), PM2.5 particles were extracted by dichloromethane and c-PAHs contents in the EOMs were determined. As markers of genotoxic potential, DNA adduct levels and oxidative DNA damage (8-oxo-7,8-dihydro-2′-deoxyguanosine, 8-oxodG, levels) induced by EOMs in an acellular assay of calf thymus DNA coupled with ³²P-postlabeling (DNA adducts) and ELISA (8-oxodG) in the presence and absence of microsomal S9 fraction were employed. Twofold higher DNA adduct levels (17.20 adducts/10⁸ nucleotides/m³ vs. 8.49 adducts/10⁸ nucleotides/m³) were induced by EOM from Ostrava-Bartovice (immediate proximity of heavy industry) compared with that from Ostrava-Poruba (mostly traffic emissions). Oxidative DNA damage induced by EOM from Ostrava-Bartovice was more than fourfold higher than damage induced by EOM from Trebon (8-oxodG/10⁸ dG/m³: 0.131 vs. 0.030 for Ostrava-Bartovice vs. Trebon, respectively). Since PM2.5 particles collected in various localities differ with respect to their c-PAHs content, and c-PAHs significantly contribute to genotoxicity (DNA adduct levels), we suggest that monitoring of PM2.5 levels is not a sufficient basis to assess genotoxicity of respirable aerosols. It seems likely that the industrial emissions prevailing in Ostrava-Bartovice represent a substantially higher genotoxic risk than mostly traffic-related emissions in Ostrava-Poruba. B[a]P and c-PAH contents in EOMs are the most important factors relating to their genotoxic potential.     

Anti-inflammatory activity of Eupatorium perfoliatum L. extracts, eupafolin, and dimeric guaianolide via iNOS inhibitory activity and modulation of inflammation-related cytokines and chemokines

Ethnopharmacological relevance

Eupatorium perfoliatum L. has been used traditionally for the treatment of fever, malaria and inflammation-associated diseases. Nowadays it is mostly used as immune activating remedy. The following study was performed to evaluate extracts with different polarity and defined lead-compounds from the herbal material on potential in vitro activities concerning immune cell activation, phagocytosis, and inflammation-related processes.

Materials and methods

MeOH-, EtOH-, and DCM extracts, beside several subfractions and isolated polysaccharides, sesquiterpene lactones and flavonoids were prepared and characterized analytically from the aerial parts of E. perfoliatum. Immunological activity was tested within lymphocyte transformation test on PBMC, test on enhancement of phagocytosis and of NO-production by murine RAW 264.7 macrophages. Anti-inflammatory effects were assessed from LPS-stimulated RAW 264.7 cells by NO/iNOS quantification, gene array, real-time PCR and ELISA.

Results

No stimulatory activity was found within lymphocyte transformation test, for phagocytic activity and NO formation in macrophages. MeOH-, EtOH- and DCM extracts showed anti-inflammatory activity against LPS-stimulated macrophages by inhibition of NO release (IC₅₀ > 100, 89, 19 μg/mL resp.) with eupafolin and a dimeric guaianolide having prominent NO inhibiting activity (IC₅₀ 6 resp. 16 μM). Anti-inflammatory activity was found on gene and protein level by significant down-regulation of cytokines CSF-3, IL-1α, IL-1β, and chemokines CCL2, CCL22 and CXCL10. Also TNF was down-regulated moderately (−17%).

Conclusions

Although the postulated immunostimulating properties of E. perfoliatum have not been confirmed, the anti-inflammatory effects can be seen as a verification of the traditional use against inflammatory diseases.