HPLC同时测定归芍疏肝散中芍药苷阿魏酸和甘草酸铵的含量

目的:建立同时测定归芍疏肝散中芍药苷、阿魏酸和甘草酸铵含量的高效液相色谱法。方法:采用Amethyst C18色谱柱(200 mm×4.6 mm,5μm),乙腈-0.1%磷酸为流动相,梯度洗脱,流速为1.0 mL·min-1,检测波长为237 nm,柱温为30℃。结果:在线性范围内3种成分线性良好(r≥0.9998),精密度RSD均<2%;重复性RSD均<3%;平均回收率98.81%~100.25%,RSD均<2%。结论:该方法快捷,操作简便,结果可靠,可用于归芍疏肝散的质量控制。     

Drosophila ammonium transporter Rh50 is required for integrity of larval muscles and neuromuscular system

Rhesus glycoproteins (Rh50) have been shown to be ammonia transporters in many species from bacteria to human. They are involved in various physiological processes including acid excretion and pH regulation. Rh50 proteins can also provide a structural link between the cytoskeleton and the plasma membranes that maintain cellular integrity. Although ammonia plays essential roles in the nervous system, in particular at glutamatergic synapses, a potential role for Rh50 proteins at synapses has not yet been investigated. To better understand the function of these proteins in vivo, we studied the unique Rh50 gene of Drosophila melanogaster, which encodes two isoforms, Rh50A and Rh50BC. We found that Drosophila Rh50A is expressed in larval muscles and enriched in the postsynaptic regions of the glutamatergic neuromuscular junctions. Rh50 inactivation by RNA interference selectively in muscle cells caused muscular atrophy in larval stages and pupal lethality. Interestingly, Rh50-deficiency in muscles specifically increased glutamate receptor subunit IIA (GluRIIA) level and the frequency of spontaneous excitatory postsynaptic potentials. Our work therefore highlights a new role for Rh50 proteins in the maintenance of Drosophila muscle architecture and synaptic physiology, which could be conserved in other species.     

Study of different routes of immunization using outer membrane vesicles of Neisseria meningitidis B and comparison of two adjuvants

Outer membrane vesicles (OMVs) of Neisseria meningitidis contain important antigens to trigger an immune response against meningococci and have been studied as vaccines compounds. The immune response to a vaccine may be affected by its constitution and route of administration. Therefore, Swiss mice were immunized by different routes with OMVs of N. meningitidis B with dimethyl dioctadecyl ammonium bromide in bilayer fragments (DDA-BF) or aluminum hydroxide (AH) as adjuvants. The adjuvants and different routes were compared regarding the immune responses by ELISA, western blot, delayed type hypersensitivity (DTH) and histopathologic analysis. The antigenic preparation generated humoral and cellular immune responses. In quantitative analyzes, in general, AH was superior to DDA-BF. However, analysis such as IgG avidity index, bactericidal activity and immunoblot, revealed no important differences regarding the adjuvant or route of immunization. Regarding the parameters tested, it was not possible to define a superiority between the adjuvants and routes of immunization proposed by this study.     

Ionic (Co)Organocatalyst with (Thio)Urea Anion and Tetra‑n‑butyl Ammonium Cation for the Polymerization of γ‐Butyrolactone

An effective ionic organocatalyst system is developed for the challenging ring‐opening polymerization (ROP) of γ‐butyrolactone (GBL) at low temperature. The catalysts are prepared by dehydration reaction between tetra‐n‐butyl ammonium hydroxide (TBAOH) and (thio)ureas at ambient temperature, and utilized with or without extra benzyl alcohol (BnOH) initiator. The solid‐state structure of TUA‐3 comprising thiourea anion is characterized by X‐ray diffraction analysis. Typically, a mixture of cyclic and linear poly(GBL) with low molecular weights (5000–1600 g mol^?1) and slightly narrow molecular distribution Ð (1.2–1.4) is obtained by single base with/without combination with (thio)ureas. Interestingly, solely linear high‐molecular‐weight poly(GBL) (10 400 g mol^?1) can be achieved by a synergistic effect of TBAOH/N,N′‐isopropylthiourea in the presence of BnOH. The obtained poly(GBL) is characterized with NMR spectroscopy and matrix‐assisted laser desorption/ionization time‐of‐flight mass spectroscopy (MALDI‐TOF MS). Mechanistic studies reveal different polymerization initiation steps in this reported catalyst system, which leads to poly(GBL) with divergent end groups.     

Dibucaine in Ionic-Gradient Liposomes: Biophysical,Toxicological, and Activity Characterization

Administration of local anesthetics is one of the most effective pain control techniques for postoperative analgesia. However, anesthetic agents easily diffuse into the injection site, limiting the time of anesthesia. One approach to prolong analgesia is to entrap local anesthetic agents in nanostructured carriers (e.g., liposomes). Here, we report that using an ammonium sulphate gradient was the best strategy to improve the encapsulation (62.6%) of dibucaine (DBC) into liposomes. Light scattering and nanotracking analyses were used to characterize vesicle properties, such as, size, polydispersity, zeta potentials, and number. In vitro kinetic experiments revealed the sustained release of DBC (50% in 7 h) from the liposomes. In addition, in vitro (3T3 cells in culture) and in vivo (zebrafish) toxicity assays revealed that ionic-gradient liposomes were able to reduce DBC cyto/cardiotoxicity and morphological changes in zebrafish larvae. Moreover, the anesthesia time attained after infiltrative administration in mice was longer with encapsulated DBC (27 h) than that with free DBC (11 h), at 320 μM (0.012%), confirming it as a promising long-acting liposome formulation for parenteral drug administration of DBC.     

Ammonium accumulation and chemokine decrease in culture media of Gcdh−/− 3D reaggregated brain cell cultures

Glutaric Aciduria type I (GA-I) is caused by mutations in the GCDH gene. Its deficiency results in accumulation of the key metabolites glutaric acid (GA) and 3-hydroxyglutaric acid (3-OHGA) in body tissues and fluids. Present knowledge on the neuropathogenesis of GA-I suggests that GA and 3-OHGA have toxic properties on the developing brain.We analyzed morphological and biochemical features of 3D brain cell aggregates issued from Gcdh^?/? mice at two different developmental stages, day-in-vitro (DIV) 8 and 14, corresponding to the neonatal period and early childhood. We also induced a metabolic stress by exposing the aggregates to 10 mM l-lysine (Lys).Significant amounts of GA and 3-OHGA were detected in Gcdh^?/? aggregates and their culture media. Ammonium was significantly increased in culture media of Gcdh^?/? aggregates at the early developmental stage. Concentrations of GA, 3-OHGA and ammonium increased significantly after exposure to Lys. Gcdh^?/? aggregates manifested morphological alterations of all brain cell types at DIV 8 while at DIV 14 they were only visible after exposure to Lys. Several chemokine levels were significantly decreased in culture media of Gcdh^?/? aggregates at DIV 14 and after exposure to Lys at DIV 8.This new in vitro model for brain damage in GA-I mimics well in vivo conditions. As seen previously in WT aggregates exposed to 3-OHGA, we confirmed a significant ammonium production by immature Gcdh^?/? brain cells. We described for the first time a decrease of chemokines in Gcdh^?/? culture media which might contribute to brain cell injury in GA-I.     

水飞蓟胶囊联合甘草酸二铵肠溶胶囊及恩替卡韦对代偿期乙肝肝硬化患者肝功能及血清T细胞亚群水平的影响

目的研究水飞蓟胶囊联合甘草酸二铵肠溶胶囊及恩替卡韦对代偿期乙肝肝硬化患者肝功能及血清T细胞亚群水平的影响。方法选取2016年1月至2018年1月本院收治的代偿期乙肝肝硬化患者90例,按照随机数字表法分为研究组和对照组,各45例。对照组采用甘草酸二铵肠溶胶囊及恩替卡韦治疗,研究组于对照组基础上加用水飞蓟胶囊治疗,两组均持续治疗6个月。比较两组乙肝病毒基因(HBV-DNA)转阴率、治疗前后肝功能指标[丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)、总胆红素(TBIL)]、中医证候积分、肝纤维化指标[血清透明质酸(HA)、层粘连蛋白(LN)、Ⅲ型前胶原(PCⅢ)、Ⅳ型胶原(C-Ⅳ)]、血清炎性因子[肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)、IL-8]及血清T细胞亚群水平变化情况。结果①HBV-DNA转阴率:研究组HBV-DNA转阴率高于对照组(P<0.05);②中医证候积分:治疗后,研究组倦怠乏力积分、纳差积分、腹胀积分、肝区不适积分、口干苦积分均低于对照组(P<0.05);③肝功能指标:治疗后,研究组血清ALT、AST、TBIL均低于对照组(P<0.05);④肝纤维化指标:治疗后,研究组血清HA、LN、PCⅢ、C-Ⅳ均低于对照组(P<0.05);⑤血清炎性因子:治疗后,研究组血清TNF-α、IL-6、IL-8均低于对照组(P<0.05);⑥血清T细胞亚群:治疗后,研究组血清CD3⁺、CD4⁺、CD4⁺/CD8⁺均高于对照组(P<0.05)。结论水飞蓟胶囊联合甘草酸二铵肠溶胶囊及恩替卡韦治疗代偿期乙肝肝硬化,能进一步缓解炎症反应,提高患者免疫功能,从而进一步抑制肝纤维化进程,改善肝功能,提高HBV-DNA转阴率,改善患者临床症状。     

基于网络药理学和设计空间的乳腺康提取工艺研究

目的通过网络药理学及设计空间法对乳腺康提取工艺参数进行研究。方法借助网络药理学对乳腺康潜在的活性成分进行筛选,并与酪氨酸激酶进行分子对接,结合《中国药典》2020年版确定指标性成分,采用HPLC法运用设计空间进行乳腺康提取工艺研究。结果筛选出乳腺康中甘草查尔酮A、川陈皮素、蒲公英甾醇等核心成分与酪氨酸激酶的亲和力与临床推荐用药相似;设计空间得到最佳提取工艺:浸泡时间为30 min、溶媒量为12倍、提取时间为45~75 min、乙醇体积分数为65%~80%、提取2~3次。结论实验所得到的工艺合理可行,验证实验与理论值预测值接近,具有一定的实用价值。该研究基于质量源于设计(QbD)理念的乳腺康提取工艺,稳定可靠,为其进一步的工艺开发及质量控制提供思路。     

Response of Human Osteoblastic and Osteoclastic Cells to AH Plus and Pulp Canal Sealer Containing Quaternary Ammonium Polyethylenimine Nanoparticles

Introduction

The incorporation of quaternary ammonium polyethylenimine (QPEI) nanoparticles into endodontic sealers induces alterations in their structure and surface properties, which may affect the compatibility with the periapical tissues. This work addressed the behavior of human bone cells exposed to extracts from commercial and QPEI containing AH Plus (DeTrey, Konstanz, Germany) and Pulp Canal Sealer EWT (PCS; Kerr Italia Srl, Salerno, Italy).

Methods

Freshly mixed AH Plus and PCS or containing 2% QPEI (0.3 mL spread over the well bottom of a 24-well plate) were extracted with culture medium (1.5 mL for 24 hours at 37°C) and diluted (1:20–1:5000). Osteoblastic or osteoclastic cells were cultured in the presence of QPEI particles (1%–10%) and were exposed to the extracts from unmodified and QPEI containing sealers.

Results

QPEI nanoparticles, at 1% and 2%, did not affect cell behavior. On osteoblastic cells, AH Plus and PCS increased DNA at 1:2500 dilution (levels ≤1:100 were cytotoxic). Alkaline phosphatase activity decreased at dilutions ≤1:500. Comparatively, QPEI containing AH Plus increased DNA at 1:2500 and 1:500 dilutions, and QPEI containing PCS induced ALP activity at 1:2500 and 1:500 dilutions. Regarding osteoclastic cells, DNA increased (AH Plus) or was not affected (PCS) at dilutions up to 1:500 and decreased with more concentrated extracts. Tartrate-resistant acid phosphatase activity decreased with dilutions ≤1:500 for both sealers. QPEI containing sealers presented a similar behavior. The sealers affected some intracellular signaling pathways, and QPEI containing sealers further modulate these mechanisms.

Conclusions

QPEI nanoparticles, at 2%, did not affect cell behavior. However, the incorporation of 2% QPEI particles into AH Plus and PCS modulates the proliferation and differentiation of bone cells, depending on the sealer and the cell type, without increasing the sealers' cytotoxicity.