Tea tree oil nanoliposomes: optimization, characterization, and antibacterial activity against Escherichia coli in vitro and in vivo.

The purpose of this study was to formulate tee tree oil nanoliposomes (TTONL) and evaluate its characterization and antibacterial activity. TTONL was prepared by thin film hydration and sonication technique, and the preparation conditions were optimized by Box-behnken response surface method. The characterization (morphology, size, zeta potential, and stability) and antibacterial activity of TTONL against Escherichia coli (E. coli) in vitro and in vivo were evaluated. The optimal preparation conditions for TTONL: lecithin to cholesterol mass ratio of 3.7:1, TTO concentration of 0.5%, and pH of the hydration medium of 7.4, which resulted in a TTONL encapsulation rate of 80.31 ± 0.56%. TTONL was nearly spherical in shape and uniform in size, and the average particle size was 227.8 ± 25.3 nm with negative charge. The specific disappearance of the TTO peak in the infrared spectrum suggested the successful preparation of TTONL, which showed high stability at 4°C within 35 d. The result of MIC test found that the nanoliposomes improved antibacterial activity of TTO against various E. coli strains. TTONL exposure in vitro caused different degrees of structural damage to the E. coli. TTONL by oral administration alleviated the clinical symptoms and intestinal lesion of chickens induced with E. coli challenge. Furthermore, TTONL treatment remarkably lowered the mRNA expression of NLRP3 and NF-κB (p65) in the duodenum and cecum of E. coli-infected chickens. In conclusion, the prepared TTONL had good stability and slow-release property with dose-dependent inhibition and killing effects on different strains of E. coli, and exerted a preventive role against chicken colibacillosis through inhibition.

Polysaccharide from Atractylodes macrocephala Koidz binding with zinc oxide nanoparticles: Characterization, immunological effect and mechanism.

Atractylodes macrocephala Koidz (A. macrocephala) has been used both as a traditional medicine and functional food for hundreds of years in Asia. And it has a variety of biological activities, such as enhancing the ability of immunity and modulating effect on gastrointestinal motility. In this study, a water-soluble polysaccharide with molecular weight of 2.743 × 103 Da was isolated from the root of A. macrocephala. Polysaccharide from A. macrocephala (AMP) consisted of arabinose, galactose, glucose, xylose, mannose, ribose, galactose uronic acid, glucose uronic acid, with a percentage ratio of 21.86, 12.28, 34.19, 0.43, 0.92, 0.85, 28.79, and 0.67%, respectively. Zinc plays an important role in immune system. Therefore, we supposed that AMP binding with zinc oxide (ZnO) nanoparticles (AMP-ZnONPs) might be an effective immunostimulator. AMP-ZnONPs was prepared by Borch reduction, and its structural features were characterized by Scanning Electron Microscope (SEM), Transmission electron microscope (TEM), TEM-energy dispersive spectroscopy mapping (TEM-EDS mapping), Fourier transform infrared spectroscopy (FT-IR), X-ray photoelectron spectrometer (XPS), X-ray diffraction (XRD), particle size and zeta-potential distribution analysis. Then, its immunostimulatory activity and the underlying mechanism were evaluated using RAW264.7 cells. The results showed that AMP-ZnONPs remarkably promoted cell proliferation, enhanced phagocytosis, the release of nitric oxide (NO), cytokines (IL-6 and IL-1ß) and the expression of co-stimulatory molecules (CD80, CD86 and MHCII). Moreover, AMP-ZnONPs could promote the expression of Toll-like receptor 4 (TLR4), Myeloid differentiation factor 88 (MyD88), TNF receptor associated factor 6 (TRAF6), phospho-IκBα (P-IκBα) and phospho-p65 (P-p65), and TLR4 inhibitor (TAK242) inhibited the expression of these proteins induced by AMP-ZnONPs. Therefore, AMP-ZnONPs activated macrophages by TLR4/MyD88/NF-κB signaling pathway, indicating that AMP-ZnONPs could act as a potential immunostimulator in medicine and functional food.

Activation of RAW264.7 macrophage by Exopolysaccharide from Aphanothece halaphytica (EPSAH) and the underlying mechanisms.

Exopolysaccharide from Aphanothece halophytica (EPSAH), a potent antitumor agent and immunological adjuvant, was investigated for the activation effect on RAW264.7 macrophages and the underlying mechanisms. EPSAH could significantly enhance macrophage phagocytosis and the secretion of nitric oxide, increase the mRNA expression levels of the pro-inflammatory cytokines (IL-1ß, IL-6, IL-12, and TNF-α), anti-inflammatory cytokine IL-10, and chemokines (MCP-1 and MIP-1α). When RAW264.7 cells were treated with EPSAH, the mRNA expression of TLR4 and its downstream molecules TRAF6 and MyD88 were upregulated. When TLR4 was blocked using a TLR4-specific neutralizing antibody, nitric oxide secretion from the macrophages was significantly inhibited. EPSAH was further shown to induce phosphorylation of the mitogen-activated protein kinases (MAPKs) ERK, JNK, and p38, and promote cytoplasmic IκB phosphorylation and increase nuclear NF-κB p65 levels remarkably in RAW264.7 cells. These data demonstrate the capacity of EPSAH to induce macrophage activation possibly via TLR4/MyD88 pathway, which leads to the activation of its main signaling downstream molecules MAPKs and NF-κB.

Comparative Studies on Chemical Contents and Effect in Kidney-Yang Deficiency Rats of Salt-Processed Product and Wine-Processed Product of Cuscutae Semen.

Cuscutae Semen mainly includes salt-processed product (SPP) and wine-processed product (WPP), which are most commonly used in traditional Chinese medicine. However, the differences between SPP and WPP have not been reported. In this paper, comparative studies between SPP and WPP on chemical contents and effect in kidney-yang deficiency rats have been investigated. UPLC-MS/MS was used to study the differences in chemical components. Kidney-yang deficiency was induced by hydrocortisone in rats. Rats were orally administrated with different doses of Jinkui Shenqi Pills, SPP, and WPP for 28 days. The average organ coefficients, renal function indexes, sex hormone levels, and testicular morphology were detected. As a result, the contents of flavonoids and chlorogenic acids were higher in SPP than in WPP, which may be caused by different processing methods. The improvement on reproduction of SPP was reflected in organ coefficients, renal function indexes, and biochemical properties of seminal plasma; furthermore, WPP was in sex hormone levels and morphology of testis. As a conclusion, these results indicated that both SPP and WPP can improve the reproductive function of kidney-yang deficiency rats with different mechanisms, which may be due to the differences in chemical contents between WPP and SPP as well as different processing methods. It is the first time that the differences between SPP and WPP have been studied in reproductive function in rats with kidney-yang deficiency.

Compounds Identification in Semen Cuscutae by Ultra-High-Performance Liquid Chromatography (UPLCs) Coupled to Electrospray Ionization Mass Spectrometry.

Semen Cuscutae is commonly used in traditional Chinese medicine and contains a series of compounds such as flavonoids, chlorogenic acids and lignans. In this study, we identified different kinds of compositions by ultra-high-performance liquid chromatography (UPLC) coupled to electrospray ionization mass spectrometry (MS). A total of 45 compounds were observed, including 20 chlorogenic acids, 23 flavonoids and 2 lignans. 23 of them are reported for the first time including 6-O-caffeoyl-ß-glucose, 3-O-(4'-O-Caffeoylglucosyl) quinic acid, etc. Their structures were established by retention behavior, extensive analyses of their MS spectra and further determined by comparison of their MS data with those reported in the literature. As chlorogenic acids and flavonoids are phenolic compounds that are predominant in Semen Cuscutae, in conclusion, phenolic compounds are the major constituents of Semen Cuscutae.

Sub-chronic toxicity study of arecae semen aqueous extract in Wistar rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Arecae semen, the ripe seed of Areca catechu L., has been used as vermifuge and digestant in traditional Chinese medicine (TCM). However, the potential toxicity effect of arecae semen has not been completely investigated. THE AIM OF THE STUDY: The present study was aimed at evaluating the sub-chronic toxicity of arecae semen by oral administration in Wistar rats. MATERIALS AND METHODS: A total of 120 Wistar rats were randomly divided into 4 groups (15 males and 15 females per group). The treated groups were given arecae semen aqueous extract (ASAE) at the dose of 750, 1500 and 4500mg/kg/day by oral administration respectively, and the control group was received distilled water only. The rats and their consumed feed were weighted every 3 days. The clinical changes and mortality were observed and recorded daily. Hematological parameters, biochemical parameters, organ weights, urinalysis and histopathological examination of all rats were tested at the end of the 30-day treatment period and another 10-day recovery period. RESULTS: Deaths, weight loss, diarrhea, sluggish action, tremors and body curl up were observed in the 1500 and 4500mg/kg groups during the study. The relative organ weights of liver and testis in male rats of 4500mg/kg group were significantly different compared with the control group at the end of the treatment period. As for laboratory parameters, there were no significant differences at the dose of 1500 and 4500mg/kg groups compared with the control group in the study, except the white blood cell count (WBC), aspartate aminotransferase (AST), alanine aminotransferase (ALT), albumin (ALB), blood urea nitrogen (BUN), creatinine (Cr), glucose (GLU) and total cholesterol (CHOL). In addition, the results of histopathological examination and feed intake showed no significant difference compared with the control group. CONCLUSIONS: The results showed that ASAE at the dose of 750mg/kg/day was safe, but long-term oral administration of ASAE with high dosage was toxic. Moreover, the toxic ingredients of ASAE including arecoline, and also some other compounds should be researched.

Identification and Analysis of Amygdalin, Neoamygdalin and Amygdalin Amide in Different Processed Bitter Almonds by HPLC-ESI-MS/MS and HPLC-DAD.

Processing is a traditional pharmacy technology based on traditional Chinese medicine theory. The traditional Chinese medicine (TCM) ingredients should be processed before being used as a medicine. Processed bitter almonds are widely used in the clinic in TCM for the treatment of cough and asthma. In this work the amygdalin profile of three producing areas in China was determined, with respect to three differently processed bitter almond products: raw, stir-fried and scalded. Identification of the compounds was done by using high performance liquid chromatography coupled to electrospray ionization mass spectrometry (HPLC-ESI-MS/MS). Results indicated that amygdalin, neoamygdalin and amygdalin amide were identified in the different processed bitter almonds. Meanwhile, amygdalin was used as a standard to calculate the quantification of amygdalin and the concentration ratio of neoamygdalin and total amygdalin by HPLC-DAD. The data suggested that composition of amygdalin isomers in bitter almonds was influenced by the processing method. It also gives a new understanding of the processing principle of bitter almonds. Moreover, the classification of different processed bitter almonds can be achieved on the basis of amygdalin isomers levels.

Purification, characterization and biological effect of reversing the kidney-yang deficiency of polysaccharides from semen cuscutae.

Semen cuscutae is a well-known Chinese medicine which has been used to nourish kidney. It is the first study to demonstrate that the polysaccharides from semen cuscutae showed significant activity of nourishing kidney-yang by increasing the levels of testosterone and estradiol, decreasing the level of blood urea nitrogen, improving immune function, possessing antioxidant effect. Three homogeneous polysaccharides were obtained by DEAE-cellulose and Sephacryl S-400 which were named as C-7WR1, C-7WR2 and C-7WR3 with average molecular weight of 7.59×104, 3.23×104 and 2.25×104 respectively. C-7WR1 was composed of fructose: mannose=0.02:1. C-7WR2 was composed of fructose: mannose: xylose: arabinose=0.01:1:0.14:0.33. C-7WR3 was composed of fructose: mannose: xylose: arabinose=0.01:1:0.10:0.47. They mainly contained mannose. Their fourier transform infrared features were similar. They all had no nucleic acid and protein.

Chemical Comparison of Two Drying Methods of Mountain Cultivated Ginseng by UPLC-QTOF-MS/MS and Multivariate Statistical Analysis.

In traditional Chinese medicine practice, drying method is an essential factor to influence the components of Chinese medicinal herbs. In this study, an ultra-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometry (UPLC-QTOF-MS/MS)-based approach was used to compare the content of chemical compounds of mountain cultivated ginseng that had been natural air dried (LX-P) and vacuum freeze-dried (LX-L). Multivariate statistical analysis such as principal component analysis (PCA) and supervised orthogonal partial least squared discrimination analysis (OPLS-DA) were used to select the influential components of different samples. There were 41 ginsenosides unambiguously identified and tentatively assigned in both LX-L and LX-P. The results showed that the characteristic components in LX-P were ginsenoside Rb1, ginsenoside Rc, ginsenoside Rg6, dendrolasin, and ginsenoside Rb2. The characteristic components in LX-L were malonyl-ginsenoside Re, malonyl-ginsenoside Rb1, malonyl-ginsenoside Rc, malonyl-ginsenoside Rb1 isomer, malonyl-ginsenoside Rb2, malonyl-ginsenoside Rb3, malonyl-ginsenoside Rd isomer, gypenoside XVII, and notoginsenoside Fe. This is the first time that the differences between LX-L and LX-P have been observed systematically at the chemistry level. It was indicated that vacuum freeze-drying method can improve the content of malonyl-ginsensides in mountain cultivated ginseng.

[Analysis of color regulation of Fluoritum in Chinese Pharmacopoeia based on the coloration mechanism of Fluorite].

The fluoritum is used for gynecology frequently and it's for those diseases: kidney yang deficiency, Gong cold sterility, palpitation due to fright, insomnia and dreaminess and cold cough. It's ruled in Chinese Pharmacopoeia (1985 edition) that the fluoritum originates from fluorite which belongs to fluoride minerals. Its main content is CaF2. The colors are of differents grades with purple or green. In the market, there are large differences in quality and it has various colors. Besides of the ruled color of purple and green, white and yellow are also common colors. By digging into and analysis the relevant research literature of fluorite which belongs to fluoride minerals, colors and coloration mechanism of fluorite are summarized in this paper.Natural fluorite is the mineral which has the most species of colors in nature. The different colors of fluorite are mainly caused by the impurity elements. At present, there are mainly about the coloration mechanism of fluorite: rare earth ions (4fN ions), color center, inclusions, crystalline domains or sub microscopic inclusions. The green of fluorite is produced by 570 nm and 305 nm absorption peaks which are caused by Sm2+ and compensated ions Na+ centers generated color center. The yellow of fluorite is produced by the joining of transition element, resulting in the formation of charge transfer between the crystal ions and the formation of O2-O32- ion molecule.The black of fluorite, mainly was attributed to the existence of a higher degree of evolution of organic matter. In this passage,suggestions for modification of the properties of fluoritum in Chinese Pharmacopoeia are put forward.

[Primary study on quality standard of carbonizing drug characteristic of ginger carbon].

OBJECTIVE: To establish the quality standard for carbonizing drug characteristic of ginger carbon. METHOD: Gingers and different carbonized gingers were compared by the absorption of pigment, tannin content, pH, mouth's coagulation time and bleeding time. RESULT: The study resulted in the recommended carbonizing standard that the absorption capacity shall not be less than 7.50 mg x g(-1) for methylene blue, the tannin content shall not be less than 2.103 mg x g(-1), the pH shall be (5.50 +/- 0.10), and coagulation time and bleeding time shall be the shorter the better. CONCLUSION: The established assessment standard for carbonizing drug characteristic of ginger carbon is reasonable, easily operated and feasible.