基于药物体系的栀子特征图谱质量表征关联分析

目的建立基于药物体系的栀子特征图谱质量表征关联分析方法与评价模式,有效精准地评价栀子饮片质量。方法运用所建立的HPLC-PDA法表征栀子药物体系特征图谱质量,基于特征图谱中特征峰的数量及所属化学类型对16批栀子饮片进行质的表征,基于特征图谱中特征指标性成分栀子苷、京尼平龙胆双糖苷和绿原酸的含量及环烯醚萜类(以栀子苷表征)、苯丙酸类(以绿原酸表征)的含量,同时采用香草醛-高氯酸显色法测定栀子总萜含量,三氯化铁-铁氰化钾显色法测定栀子总酚含量,对16批栀子饮片进行量的表征,并基于基准饮片将16批栀子饮片质与量的表征结果分别进行关联性分析。结果以批号6为基准,栀子饮片特征图谱中共含有特征峰9个,其中环烯醚萜成分特征峰4个,苯丙酸类成分特征峰5个,16批饮片色谱图中均含有此9个特征峰。批号3、4、7、8、13有效指标性成分总体含量高于基准批号6,批号7、1、9、4、3、12、5、14与基准批号6关联性最高,综合评价得出批号3、4、6、7优良度居前。结论建立了栀子饮片的HPLC特征图谱质量表征方法,并结合总酚及总萜含量测定对16批栀子饮片进行质量表征。所建立的基于药物体系的栀子特征图谱质量表征关联分析模式,关注了栀子饮片质量整体关联性与应用有效性,能有效精准地评价栀子饮片质量。     

Design,synthesis, and evaluation of genipin derivatives for the treatment of Alzheimer's Disease

Twenty‐two novel genipin derivatives have been designed, synthesized, and evaluated for their inhibitory activity against acetylcholinesterase (AChE). As a result, compound 13a bearing ligustrazine moiety displayed the most potent AChE inhibitory activity in this series with IC₅₀ value of 218 nm . Besides, MTT assay was performed to investigate the neuroprotection of these compounds against PC12 cells injured by Amyloid β‐protein 1–42 (Aβ_1–42). Among them, 8a showed higher inhibition rate (%Inhibition = 22.29) than the positive reference Donepezil (%Inhibition = 17.65).     

Exogenous collagen cross-linking recovers tendon functional integrity in an experimental model of partial tear

We investigated the hypothesis that exogenous collagen cross-linking can augment intact regions of tendon to mitigate mechanical propagation of partial tears. We first screened the low toxicity collagen cross-linkers genipin, methylglyoxal and ultra-violet (UV) light for their ability to augment tendon stiffness and failure load in rat tail tendon fascicles (RTTF). We then investigated cross-linking effects in load bearing equine superficial digital flexor tendons (SDFT). Data indicated that all three cross-linking agents augmented RTTF mechanical properties but reduced native viscoelasticity. In contrast to effects observed in fascicles, methylglyoxal treatment of SDFT detrimentally affected tendon mechanical integrity, and in the case of UV did not alter tendon mechanics. As in the RTTF experiments, genipin cross-linking of SDFT resulted in increased stiffness, higher failure loads and reduced viscoelasticity. Based on this result we assessed the efficacy of genipin in arresting tendon tear propagation in cyclic loading to failure. Genipin cross-linking secondary to a mid-substance biopsy-punch significantly reduced tissue strains, increased elastic modulus and increased resistance to fatigue failure. We conclude that genipin cross-linking of injured tendons holds potential for arresting tendon tear progression, and that implications of the treatment on matrix remodeling in living tendons should now be investigated.     

Genipin Inhibits RANKL-Induced Osteoclast Differentiation Through Proteasome-Mediated Degradation of c-Fos Protein and Suppression of NF-κB Activation

People over the age of 50 are at risk of osteoporotic fracture, which may lead to increased morbidity and mortality. Osteoclasts are responsible for bone resorption in bone-related disorders. Genipin is a well-known geniposide aglycon derived from Gardenia jasminoides, which has long been used in oriental medicine for controlling diverse conditions such as inflammation and infection. We aimed to evaluate the effects of genipin on RANKL-induced osteoclast differentiation and its mechanism of action. Genipin dose-dependently inhibited early stage RANKL-induced osteoclast differentiation in bone marrow macrophages (BMMs) during culture. Genipin inhibited RANKL-induced IκB degradation and suppressed the mRNA expression of osteoclastic markers such as NFATc1, TRAP, and OSCAR in RANKL-treated BMMs, but did not affect c-Fos mRNA expression. Interestingly, genipin markedly inhibited c-Fos protein expression in BMMs, which was reversed in the presence of the proteosome inhibitor MG-132. Furthermore, genipin inhibited RANKL-mediated osteoclast differentiation, which was also rescued by overexpression of c-Fos and NFATc1 in BMMs. Taken together, our findings indicate that genipin down-regulated RANKL-induced osteoclast differentiation through inhibition of c-Fos protein proteolysis as well as inhibition of IκB degradation. Our findings indicate that genipin could be a useful drug candidate that lacks toxic side effects for the treatment of osteoporosis.     

京尼平-β-环糊精包合物制备工艺研究

 目的研究京尼平-β-环糊精包合物最佳制备工艺。方法采用研磨法,以包合物的含量和京尼平(genipin,GP)收率为考察指标,通过正交实验设计,综合平衡后确定最佳制备工艺。结果最佳包合条件为:包合时间1 h、GP和β-环糊精的配比(摩尔比)1∶1、包合溶剂为水。经红外光谱法、紫外光谱法、电镜扫描法、X-射线衍射法和相溶解度法鉴定,确证形成京尼平-β-环糊精包合物。结论采用最优包合条件制备的包合物收率为76.27%,京尼平含量为16.5 mg·g^-1。制成包合物后提高了京尼平的稳定性,制备工艺简单,同时也为京尼平制备成各种剂型开辟良好的前景。     

栀子及其提取物对大白鼠胆胰功能的影响

用胆胰插管研究了中药栀子及其不同提取物对大鼠胆胰流量及胰酶活性的影响。结果表明,栀子及其几种提取物有明显的利胰、利胆及降胰酶效应,但不同浓度、不同品种及不同提取物在胆胰作用上存在一定差别。京尼平甙有最显著的降低胰淀粉酶作用,而其酶解物京尼平的增加胰胆流量作用最强,持续时间较短。结合到中药栀子具有抑菌消炎及能引起胆囊收缩,降低胆道括约肌紧张性的作用,可以认为栀子可望成为急性胰腺炎及胆道感染的有价值的治疗药物。     

Comparison of natural crosslinking agents for the stabilization of xenogenic articular cartilage

Osteochondral xenografts are potentially inexpensive, widely available alternatives to fresh allografts. However, antigen removal from xenogenic cartilage may damage the extracellular matrix and reduce compressive stiffness. Non‐crosslinked xenogenic cartilage may also undergo rapid enzymatic degradation in vivo. We hypothesized that natural crosslinking agents could be used in place of glutaraldehyde to improve the mechanical properties and enzymatic resistance of decellularized cartilage. This study compared the effects of genipin (GNP), proanthocyanidin (PA), and epigallocatechin gallate (EGCG), on the physical and mechanical properties of decellularized porcine cartilage. Glutaraldehyde (GA) served as a positive control. Porcine articular cartilage discs were decellularized in 2% sodium dodecyl sulfate and DNase I followed by fixation in 0.25% GNP, 0.25% PA, 0.25% EGCG, or 2.5% GA. Decellularization decreased DNA by 15% and GAG by 35%. For natural crosslinkers, the average degree of crosslinking ranged from approximately 50% (EGCG) to 78% (GNP), as compared to 83% for the GA control. Among the natural crosslinkers, only GNP significantly affected the disc diameter, and shrinkage was under 2%. GA fixation had no significant effect on disc diameter. Decellularization decreased aggregate modulus; GA and GNP, but not EGCG and PA, were able to restore it to its original level. GNP, PA, and GA conferred a similar, almost complete resistance to collagenase degradation. EGCG also conferred substantial resistance but to a lesser degree. Overall, the data support our hypothesis and suggest that natural crosslinkers may be suitable alternatives to glutaraldehyde for stabilization of decellularized cartilage. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1037–1046, 2016.     

基于网络药理学的黄芩素、京尼平抗脑缺血作用机制研究

目的采用网络药理学及分子对接技术,以灯盏花乙素苷元(scutellarein,SE)作为参照,预测黄芩素(baicalein,BE)、京尼平(genipin,GE)抗脑缺血作用靶点,为脑缺血疾病的临床防治和开发研究提供参考。方法利用TCMSP、Swiss Target Prediction和Stitch数据库检索和文献挖掘方法,预测SE、BE、GE作用靶点,同时应用DisGeNET、CTD、NCBI Gene、OMIM、DrugBank和PharmGkb数据库预测脑缺血疾病相关靶点。通过Cytoscape 3.3.0构建小分子-靶点网络图,用DAVID软件分析SE、BE、GE特有抗脑缺血靶点的GO功能富集和KEGG通路分析。采用AutodockVina软件对SE、BE、GE与抗脑缺血共同靶点分别进行对接研究,根据受体与配体的结合能和抑制浓度排序,选择最佳的靶点蛋白。结果 SE、BE、GE分别预测出30、59、35个靶蛋白,SE与BE共同抗脑缺血疾病靶点有PIK3CG、CYP1A2、VEGFA、ALOX5、PTGS2,SE与GE共同抗脑缺血靶点为PTGS2。分子对接结果显示受体PTGS2与SE、BE、GE对接结合能及抑制浓度相对较低。GO功能富集结果显示BE-SE共同靶点主要分布于细胞质、细胞器膜、内质网膜等部位,具有与金属离子、阳离子结合功能及催化、氧化还原酶活性,参与细胞脂质、羧酸、含氧酸、有机酸代谢及脂肪酸合成等过程有关;KEGG分析结果显示受体PTGS2主要作用于花生四烯酸代谢通路和VEGF信号通路,BE与GE配伍治疗脑缺血疾病通过抑制PTGS2(COX-2)和VEGF蛋白的表达,从而减轻炎症因子造成的脑组织损伤并改善血脑屏障(Blood brain barrier,BBB)的通透性发挥脑保护作用。结论预测了BE与GE配伍治疗脑缺血疾病的潜在靶点,初步验证了治疗脑缺血疾病的作用机制,为进一步深入研究BE与GE配伍治疗脑缺血作用机制提供参考,也为下一步合成新的衍生物提供基础。     

Fabrication and characterization of Chinese giant salamander skin composite collagen sponge as a high-strength rapid hemostatic material

Chinese giant salamander (CGS) has high medicinal value and long history of clinical use in ancient China. In this study, CGS skin (CGSS) collagen was extracted and purified to prepare collagen sponge by freeze-drying. TEMPO oxidized microfibrillated cellulose (TEMPO-MFC) and genipin were adopted to improve the mechanical properties of collagen sponge. The hygroscopicity, porosity, mechanical properties, hemostatic performance, morphology, and biodegradability of the resultant sponges were investigated in detail. The results indicated that CGSS collagen was type I collagen with intact triple-helical structure, and the prepared sponge had porous structure and excellent hemostatic performance with procoagulant ratio of 53.28%. However, the CGSS collagen sponge showed low tensile strength (TS) of 98.80?KPa, compression strength (CS) of 1.48?KPa, and elongation at break (E) of 4.72%. Incorporating 2.5% TEMPO-MFC into the native CGSS collagen sponge resulted in an increase of 188.26% in TS to 284.80?KPa, 166.89% in CS to 3.95?KPa, and 73.52% in E to 8.19%. The improvements were attributed to the physical filling of TEMPO-MFC in cavity and cavity wall of collagen sponge and the stable chemical linkage between carboxyl of TEMPO-MFC and amino group of collagen which effectively improved the toughening of sponge and formed good interface bonding, respectively. Subsequent 0.3% genipin treatment further improved the TS to 605.00?KPa and the CS to 8.66?KPa as a result of crosslinking reaction. Moreover, the composite reinforcing also improved the anti-degradation ability and procoagulant ratio of collagen sponge. All results suggested that the TEMPO-MFC toughened and genipin crosslinked CGSS composite collagen sponge is a promising rapid hemostatic material with high-strength and can be applicated in biomedical field.     

Crosslinking Efficacy and Cytotoxicity of Genipin and Its Activated Form Prepared by Warming It in a Phosphate Buffer: A Comparative Study

The aim of the present study was to compare the acute and cumulative cytotoxicity of intact (n-GE) and warmed genipin (w-GE), while investigating the differences in crosslinking capabilities of these two genipins by rheological and mechanical tests. The n-GE solution was prepared by dissolving genipin powder in a sodium phosphate buffer solution. The w-GE solution was prepared by warming the n-GE solution at 37 °C for 24 h. The mechanical tests for chitosan (CH)/genipin gels showed the crosslinking rate of w-GE was much greater than that of n-GE up until 6 h after preparation, whereas the degree of crosslinking of CH/n-GE gels became higher at 12 h. The ISO 10993-5 standard method, which is established specifically for evaluating cumulative cytotoxicity, determined equivalent IC50 for w-GE (0.173 mM) and n-GE (0.166 mM). On the other hand, custom-made cytotoxicity tests using a WST-8 assay after 1 h of cultivation showed that the acute cytotoxicity of w-GE was significantly higher than that of n-GE at concentrations between 0.1–5 mM. The acute cytotoxicity of w-GE should be taken into consideration in its practical uses, despite the fact that the much faster crosslinking of w-GE is useful as an effective cross linker for in-situ forming gels.