The incidence rate of cognitive dysfunction (CD) urgently requires the development of safe and effective dietary supplements to prevent and treat this disease. Polygonatum cyrtonema Hua (PS) is a traditional Chinese herb that can be used as both food and a medicine, and is known to improve memory and nourish the nerves. However, there has been limited research on the commonly used wine-processed form of PS (WPS) in clinical practice; consequently, we know little about the pharmacological effects of WPS and the mechanisms involved. In this study, we investigated the homogeneous polysaccharides extracted from PS (SP3) and WPS (JP3) and investigated the effects of JP3 on the CD and the brain-gut axis in chronic unpredictable mild stress (CUMS) mice. SP3 and JP3 showed significant differences in terms of molecular weight, monosaccharide content, and surface morphology. Mechanistic studies showed that JP3 affected the brain-gut axis by regulating the gut microbiota and the intestinal barrier, and improved lesions and oxidative stress in the hippocampus, thus alleviating cognitive impairment. JP3 was able to increase probiotics such as Lactobacillus murinus, improve hippocampal lesions, and oxidative stress levels. Compared with inulin, JP3 better promotes the proliferation of Lactobacillus murinus and the production of butyric acid and isobutyric acid. The results suggest that WPS homogeneous polysaccharides can regulate intestinal microbiota and improve cognitive level, which is a potential functional product for the prevention of CD.

Polygonatum cyrtonema Hua (PC) is a traditional Chinese medicine with a long history of use as pharmaceuticals or functional foods. Wine steaming is the main processing method of PC, which changes the structure of polysaccharides and oligosaccharides in PC and enhances biological activities. This study investigated the structural characterization of raw and wine-steamed PC oligosaccharides and the differences in the immunomodulatory effects using cyclophosphamide (CTX)-induced immunosuppression rat model. The oligosaccharides content and molecular weight of PC after wine steaming decreased, the proportion of oligosaccharides in total sugars and the reducing sugars content increased, and the monosaccharides composition of oligosaccharides changed. The raw Polygonatum cyrtonema Hua oligosaccharides (PCCO) and the wine-steamed Polygonatum cyrtonema Hua oligosaccharides (PCWO) exerted regulatory effects on organ index, immunoglobulin G (IgG), complement 3 (C3) spleen and colon tissue morphology, hematopoietic function of immunosuppressive rats treated by cyclophosphamide (CTX). Both PCCO and PCWO significantly regulated and improved the diversity and abundance of gut microbiota in immunosuppressed rats and increased the content of short-chain fatty acids (SCFAs) in the feces of rats, and the regulating effect of PCWO was better than PCCO. Differential microbiota analysis showed that PCWO could promote the proliferation of Bifidobacterium, Bacteroides, Oscillibacter, Roseburia, and Alistipes. In summary, the difference in the structural characteristics of PC oligosaccharides might be the reason for immune enhancement. This study could provide a theoretical basis for clarifying the scientific connotation of wine steaming to enhance the efficacy of PC, and promote the application of wine-steamed PC as an immunomodulator in pharmaceuticals or functional foods.

Euphorbia lathyris L. (EL) is a traditional poisonous herbal medicine used to treat dropsy, ascites, amenorrhea, anuria and constipation. Processing to reduce toxicity of EL is essential for its safe and effective application. However, there is little known regarding the molecular mechanism of reducing toxicity after EL processing. This research aimed to screen the differential markers for EL and PEL, explore the differential mechanisms of inflammatory injury induced by EL and processed EL (PEL) to expound the mechanism of alleviating toxicity after EL processing. The results showed that 15 potential biomarkers, mainly belonging to diterpenoids, were screened to distinguish EL from PEL. EL promoted the expressions of TLR4, NLRP3, NF-κB p65, IL-1β and TNF-α, increased lipid rafts abundance and promoted TLR4 positioning to lipid rafts. Meanwhile, EL decreased LXRα and ABCA1 expression, and reduced cholesterol efflux. In contrast to EL, the effects of PEL on these indicators were markedly weakened. In addition, Euphorbia factors L1, L2, and L3 affected LXRα, ABCA1, TLR4, NLRP3, NF-κB p65, TNF-α and IL-1β expression, influenced cholesterol efflux and lipid rafts abundance, and interfered with the colocalization of TLR4 and lipid rafts. The inflammatory injury caused by processed EL was significantly weaker than that caused by crude EL, and reduction of Euphorbia factors L1, L2, and L3 as well as attenuation of inflammatory injury participated in processing-based detoxification of EL. Our results provide valuable insights into the attenuated mechanism of EL processing and will guide future research on the processing mechanism of toxic traditional Chinese medicine.

目的 探讨延胡索总碱贴片的安全性及其对神经病理性疼痛大鼠的镇痛作用及其机制.方法 ①安全性实验:采用单次、多次给药的皮肤刺激性实验及皮肤过敏性实验,评价延胡索总碱贴片的安全性.②镇痛实验:取48 只雄性SD大鼠,随机分为正常组、假手术组、模型组、布洛芬组、双氯芬酸钠贴片组及延胡索总碱贴片高、中、低剂量组,每组 6 只.除正常组和假手术组外,其余组大鼠采用坐骨神经慢性压迫损伤方法诱导神经病理性疼痛模型,术后第8 天开始,布洛芬组灌胃给药,双氯芬酸钠贴片组及延胡索总碱贴片高、中、低剂量组分别给予双氯芬酸钠贴片及延胡索总碱贴片腹部贴敷,其余组腹部贴敷空白贴片,均每天1 次,连续14 d.分别于术前及干预后1,3,7,10,14 d测定各组大鼠热缩足反射潜伏期;采用Western blot法检测各组大鼠干预结束后脊髓中Toll样受体4(TLR4)、核因子κB(NF-κB)、p38 丝裂原活化蛋白激酶(p38MAPK)、信号传导和转录激活因子3(STAT3)蛋白表达情况.结果 ①给予延胡索总碱贴片后,皮肤刺激反应评分均小于0.5 分,致敏率为0,组织结构与正常皮肤无差异.②镇痛实验中,延胡索总碱贴片中、高剂量组干预后3 d和延胡索总碱贴片各剂量组干预后7,10,14 d患侧的热缩足反射潜伏期均明显长于同期模型组(P均<0.05);干预结束后,延胡索总碱贴片各剂量组大鼠脊髓中 TLR4、NF-κB、p38MAPK蛋白相对表达量均明显低于模型组(P均<0.05),STAT3 蛋白相对表达量均明显高于模型组(P均<0.05).结论 延胡索总碱贴片安全性良好,可抑制慢性坐骨神经挤压损伤神经病理性疼痛大鼠痛觉敏感,其机制可能与抑制脊髓中TLR4、NF-κB、p38MAPK表达并上调STAT3 的表达有关.

Objective Using the stable isotope ratio method for the authenticity identification and variety identification of refined honey. Methods In this paper, a total of 17 samples of different varieties of refined honey were used to obtain refined honey proteins by precipitation with sodium tungstate solution and sulphuric acid solution. The isotope mass spectrometer was used to simultaneously detect the δ 13 C values of refined honey proteins and refined honey as well as the δ 18 O and δ 2 H values of refined honey, processed of the results obtained, analysed the authenticity of the samples and conduct a variety identification study. Results Tested of the resulting honey samples, the results showed that four batches of refined honey did not up to standard, two batches of C-4 vegetable syrup were detected as adulterated, and two batches of protein were not detected. The δ 18 O and δ 2 H values of refined honey were also found to be effective in distinguishing the varietal origin of refined honey to a certain extent. Conclusions The stable isotope ratio method is useful in the authenticity identification of refined honey, and provides new ideas to further promote the authenticity of refined honey and variety identification research.

Objective To study the mechanism of extracts from Qianjinzi(Euphorbiae Semen)and Qianjinzishuang(Euphorbiae Semen Pulveratum)mediating Toll-like receptor 4(TLR4)signaling pathway in Caco-2 cells via regulating lipid rafts.Methods Caco-2 cells were divided into Euphorbiae Semen low-,medium-and high-dose(50,200,800 μg/mL)groups,Euphorbiae Semen Pulveratum low-,medium-and high-dose(50,200,800 μg/mL)groups and control group.Flow cytometry was used to detect the lipid rafts abundance of Caco-2 cells membrane.Confocallaser scanning microscope was used for studying the co-localization of TLR4 and lipid rafts in Caco-2 cells.Western blotting was applied to detect TLR4,nuclear factor-κB p65(NF-κB p65),p-NF-κB p65 and NOD-like receptor family pyrin domain containing 3(NLRP3)protein expressions in Caco-2 cells.Furthermore,ELISA was applied to determine the levels of interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)in the supernatant of Caco-2 cells.And the levels of IL-1β and TNF-α were detected in Caco-2 cells after transfection with TLR4.Results Compared with control group,Euphorbiae Semen extracts could increase the content of lipid rafts(P＜0.01),promote the recruitment of TLR4 to lipid rafts(P＜0.01),increase the protein expressions of TLR4,NF-κB p65,p-NF-κB p65 and NLRP3,as well as induce the release of IL-1β and TNF-α in Caco-2 cells(P＜0.05,0.01,0.001).However,the effects of Euphorbiae Semen Pulveratum extracts on the abundance of lipid rafts,the recruitment of TLR4 to lipid rafts,the conduction of TLR4-mediated signaling pathways and the secretion of pro-inflammatory factors were significantly weaker than those of Euphorbiae Semen extracts at the same dose(P＜0.05,0.01,0.001).In addition,the results of siTLR4 transfection in vitro further confirmed that the content of IL-1β and TNF-α were significantly decreased after transfection of TLR4 in Caco-2 cells(P＜0.05,0.01,0.001).Conclusion The attenuation mechanism of processing for Euphorbiae Semen may be closely related to interfering the recruitment of TLR4 to lipid rafts by regulating lipid rafts abundance,and ultimately affecting the cascade of TLR4 signaling protein contained in lipid rafts.

目的 优化酒黄精wine-processed Polygonati Rhizoma的高压蒸制工艺,确定最佳工艺参数,并对酒制前后刺激性、免疫调节及降血糖作用进行研究.方法 以润制时间、蒸制压力、蒸制时间和焖制时间作为考察因素,以性状、黄精多糖、薯蓣皂苷元、水浸出物、醇浸出物和5-羟甲基糠醛(5-hydroxymethylfurfural,5-HMF)作为考察指标,运用AHP-熵权法结合 Box-Behnken 设计-响应面法(Box Behnken design-response surface methodology,BBD-RSM)确定酒黄精炮制工艺条件参数.通过家兔眼刺激性实验,评价黄精酒制前后的刺激性作用.通过ip环磷酰胺建立小鼠免疫抑制模型,比较黄精酒制前后对免疫调节作用的影响.通过饲喂高脂高糖饲料、ip链脲佐菌素建立糖尿病小鼠模型,比较黄精酒制前后对降血糖作用的影响.结果 酒黄精炮制最佳工艺参数为润制时间5 h、蒸制时间1h、蒸制压力0.06 MPa、焖制时间7 h.药效实验结果表明,生黄精具有刺激性,黄精酒制后对黏膜的刺激作用减弱,酒黄精提高小鼠免疫力和降血糖的作用均强于生黄精.结论 优选出的酒黄精炮制工艺稳定、合理、可行,按照此工艺制备的酒黄精,可以降低其刺激性,增强免疫调节作用和降血糖作用,为黄精的炮制工艺研究和临床用药提供参考.

目的 研究延胡索总碱贴片中延胡索总碱在不同条件下的稳定性.方法 以延胡索总碱贴片中的延胡索甲素、延胡索乙素以及延胡索丑素的含量,24h体外累积释放量及黏附力为指标,进行了强光照射、高温、高湿的影响因素试验、加速试验及长期留样试验.结果 制剂对光照、高温、高湿稳定,加速试验、长期留样试验稳定性较好,延胡索总碱贴片中的延胡索甲素、延胡索乙素和延胡索丑素的含量、24h体外累计释放量及延胡索总碱贴片黏附力无显著性变化.结论 延胡索总碱贴片质量可控,稳定性良好.

Objective To investigate the effects of the extracts from Semen Euphorbiae before and after frosting on the toll-like receptor 4 （TLR4）/nuclear factor kappa-B （NF-κB）/nod-like receptor family pyrin domain containing 3 （NLRP3） signaling pathway. Methods According to the body weight， 42 BALB/c mice were randomly divided into a blank control group， Semen Euphorbiae low-， medium-， and high-dose groups （10.49， 20.98， and 41.96 g/kg， respectively） and Semen Euphorbiae Pulveratum low-， medium-， and high-dose groups （10.49， 20.98， and 41.96 g/kg， respectively）， with six rats per group. Mice received continuous gavage for 7 d. The contents of interleukin-1β （IL-1β） and tumor necrosis factor-α （TNF-α） in serum of mice were determined by enzyme linked immune sorbent assay. The protein and the mRNA expression of TLR4， NF-κB p65， and NLRP3 in colon tissues of mice were detected by Western blotting and real-time PCR. According to the body weight， 30 BALB/c mice were randomly divided into a blank control group， Semen Euphorbiae group， Semen Euphorbiae+TAK-242 group， Semen Euphorbiae Pulveratum group， and Semen Euphorbiae Pulveratum+ TAK-242 group， with six rats per group. The administration groups were respectively given 20.96 g/kg of Semen Euphorbiae or Semen Euphorbiae Pulveratum， and the continuous gavage was 7 d. The Semen Euphorbiae+TAK-242 group and Semen Euphorbiae Pulveratum+ TAK-242 group were intraperitoneally injected with TAK242 （3 mg/kg） 30 min before and on days 5， 6， and 7 of administration. The protein expression of NLRP3 and NF-κB p65， and the mRNA expression of NLRP3 and IL-1β in colon tissues of mice were assessed by Western blotting and real-time PCR. Results Semen Euphorbiae not only significantly promoted the release of IL-1β and TNF-α， but it also significantly increased the protein and the mRNA expression of TLR4， NF-κB p65 and NLRP3 （P ＜ 0.05， P ＜ 0.01）. However， upregulation of IL-1β and TNF-α secretion， and TLR4， NF-κB p65， NLRP3 protein， and mRNA expression were decreased by Semen Euphorbiae Pulveratum （P ＜ 0.05， P ＜ 0.01）. NLRP3 and NF-κB p65 protein expression and NLRP3 and IL-1β mRNA expression were inhibited after the intervention with the TLR4 inhibitor TAK-242 （P ＜ 0.01）. Conclusion Semen Euphorbiae could induce an inflammatory response of the colon tissue by activating the TLR4-mediated NF-κB/NLRP3 pathway， thus promoting the release of TNF-α and IL-1β. However， upregulation of the TLR4/NF-κB/NLRP3 signaling pathway and pro-inflammatory factor secretion were significantly alleviated by Semen Euphorbiae Pulveratum， indicating that the detoxification mechanism of Semen Euphorbiae frosting may be related to the decreased ability to cause inflammation and the intervention of TLR4/NF-κB/NLRP3 signaling pathway.

目的 运用网络药理学和分子对接探讨千金子二萜醇酯类成分治疗白血病的潜在作用机制.方法 选取千金子中7种代表性二萜醇酯类成分为研究对象,综合运用SwissTargetPre-diction、SEA、OMIM、Gene Cards数据库筛选及预测千金子二萜醇酯类成分治疗白血病的潜在靶点.通过STRING数据库和Cytoscape 3.7.2软件来建立PPI网络.使用DAVID数据库对筛选出的靶点进行基因本体(GO)生物学过程分析和京都基因与基因组百科全书(KEGG)通路分析,并应用Cytoscape 3.7.2构建"成分-靶点-通路"网络,最后对千金子二萜醇酯类成分与关键靶蛋白进行分子对接验证.结果 7种千金子二萜醇酯类成分涉及252个作用靶点,筛选得到75个治疗白血病的潜在靶点,PPI核心网络表明治疗白血病的关键靶点为CDK1、CDK2、CCNA2,基因富集分析及"成分-靶点-通路"网络图显示千金子二萜醇酯类成分可能通过调节PI3 K-Akt、ErbB、p53等信号通路来发挥对白血病的治疗作用.结论 本研究表明千金子二萜醇酯类成分可能通过调节细胞凋亡、增殖分化、转录、细胞周期等多种途径来治疗白血病,诠释了中药通过多成分、多靶点、多途径治疗疾病的研究思路,为进一步深入研究千金子二萜醇酯类成分治疗白血病的作用机制提供了科学依据.

The purpose of this study was to evaluate the pharmacokinetics and tissue distribution of Corydalis yanhusuo total alkaloids transdermal patch (CTTP) following Shenque acupoint application in rats. The concentrations of Corydaline (CDL), Tetrahydropalmatine (THP), Tetrahydrocolumbamine (THC), Protopine (PRO), and Dehydrocorydaline (DHC) in rat plasma and various tissues were simultaneously detected by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) after Shenque acupoint administration of CTTP. Plasma, heart, liver, spleen, lung, and kidney tissue samples were collected at specific times and separated by gradient elution on an ACQUITY UPLC HSS T3 column (1.8 μm, 100 mm×2.1 mm) with a mobile phase of 0.01% formic acid aqueous solution and acetonitrile-0.01% formic acid. The methodological results showed that the selectivity, linear range, accuracy, precision, stability, matrix effect, and extraction recovery of the established method met the requirements of biological sample analysis. The results denoted that CTTP following Shenque acupoint administration rapidly delivered adequate drug into rat blood and maintained an effective plasma level for a significantly longer time than non-acupoint administration. Furthermore, CTTP effectively reached the liver through Shenque acupoint administration and showed tissue selectivity. The data obtained could provide a prospect for the treatment of chronic pain with CTTP following Shenque acupoint application.

以千金子中3种主要的二萜醇酯类成分提取率的综合评分为评价指标,通过Box-Behnken响应面法结合熵权法对千金子二萜醇酯类成分的闪式提取工艺进行优化.确定最佳提取工艺为:料液比1:14(g:mL)、提取时间2.5 min、提取电压130 V,在此条件下,3种二萜醇酯类成分提取率的综合评分为0.370,与模拟预测值(0.396)接近.该闪式提取工艺设计合理,可为千金子主要二萜醇酯类成分的快速高效提取提供一定的参考.

目的:研究甘草炮制前后对免疫低下小鼠免疫调节作用的影响.方法:小鼠腹腔注射环磷酰胺制备免疫低下模型,给予生甘草、清炒甘草、蜜炙甘草的提取液,检测免疫功能学评价指标,包括免疫器官指数、半数溶血值、溶血空斑数、脾淋巴细胞增值率、自然杀伤(NK)细胞杀伤率、耳肿胀度、碳廓清指数、腹腔巨噬细胞吞噬率及吞噬指数.结果:与对照组小鼠比较,模型组小鼠各检测指标明显降低(P<0.05,P<0.01);与模型组小鼠比较,各给药组(生甘草组、清炒甘草组、蜜炙甘草组)小鼠各检测指标均能增强.其中,仅蜜炙甘草组能明显增强各检测指标(P<0.05,P<0.01);与生甘草组小鼠相比,清炒甘草组能明显增强血清溶血素和抗体生成细胞、脾淋巴细胞增殖率和NK细胞活性(P<0.05,P<0.01),但对胸腺指数、脾脏指数、腹腔巨噬细胞吞噬率和吞噬指数、DTH、碳廓清功能无明显影响,蜜炙甘草组能明显增强除胸腺指数的各项评价指标(P<0.05,P<0.01),但对胸腺指数无明显影响.结论:甘草蜜炙前后对小鼠非特异性免疫、体液免疫及细胞免疫功能均有促进作用,且蜜炙后作用更强,为蜜炙甘草机制研究提供参考.

Previous pharmacological studies have indicated that diterpenoids are the primary effective chemical cluster in the seeds of Euphorbia lathyris L. The seed products are used in traditional Chinese medicine in the forms of Semen Euphorbiae (SE) and Semen Euphorbiae Pulveratum (SEP). However, the metabolism of the plant's diterpenoids has not been well elucidated, which means that the in vivo metabolite products have not been identified. The current study screened the physiological metabolites of six diterpenes [Euphorbia factor L1 (L1), L2 (L2), L3 (L3), L7a (L7a), L7b (L7b), and L8 (L8)] in feces and urine of rats after oral administration of SE and SEP using UHPLC-Q-Exactive MS. A total of 22 metabolites were detected in feces and 8 in urine, indicating that the major elimination route of diterpenoids is via the colon. Hydrolysis, methylation, and glucuronidation served as the primary metabolic pathways of these diterpenoids. In sum, this study contributed to the elucidation of new metabolites and metabolic pathways of SE and SEP, and the new chemical identities can be used to guide further pharmacokinetic studies.

目的 考察延胡索总碱的溶解度、油水分配系数等理化性质,研究促透剂及穴位、非穴位贴敷对其体外经皮渗透特性的影响,为指导其经皮给药剂型设计和制备提供参考.方法 建立酸性染料比色法和高效液相色谱(high performance liquid chromatography,HPLC)法对延胡索总碱、延胡索甲素、延胡索乙素及延胡索丑素的含量进行测定,应用饱和溶液法和经典摇瓶法检测延胡索总碱的溶解性及油水分配情况;采用Franz扩散池法考察大鼠神阙穴与非穴位皮肤体外经皮渗透性能.结果 结果显示,延胡索总碱纯度大于50％,其在10％乙醇的生理盐水中溶解性较好,可作为扩散池接收介质.当pH在5.5～7.2时,延胡索总碱的logP值控制在-1～0之间,说明延胡索总碱为强亲水性组分,延胡索甲素、延胡索乙素、延胡索丑素的logP值均在0～3之间,体外经皮渗透性良好,神阙穴用药组的24小时体外累积透过量和滞留量优于非穴位(P<0.05).结论 延胡索总碱具有适宜经皮渗透的理化性质及较好的经皮渗透性能,适宜制成经皮给药制剂,且经神阙穴用药具有一定优势,以上结果可为经皮给药剂型设计提供实验基础.