In Vitro Metabolite Profiling and Anti-Inflammatory Activities of Rhodomyrtus Tomentosa with Red Blood Cell Membrane Stabilization Methods

Background:Rhodomyrtus Tomentosa (Karamunting) is one of South Kalimantan''s biodiversity. In previous research on asthma and coal dust exposure models, nebulization with an ethanol extract of R. tomentosa leaves has been shown to reduce inflammatory cells. This study aimed to investigate the anti-inflammatory activity on the stabilization of red blood cell membranes and to identify the chemical compounds present in extracts of R. tomentosa leaves.Methods:Anti-inflammatory effect of R. tomentosa leaves was evaluated by in vitro red blood cell membrane stabilization methods. Nonsteroidal anti-inflammatory sodium diclofenac was used as the reference drug. The content of chemical compounds in the ethanol extract of R. tomentosa leaves was identified using Ultra Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS).Results:The results of inhibition of red blood cells membrane lysis showed the n-hexane fraction (concentration 25 ppm), ethyl acetate fraction (concentration 50 ppm), and a fraction of water (concentration 50 ppm) with an inhibition level of 54.5%, 81.8%, 63.6%, respectively. The ethyl acetate fraction showed the highest inhibition of hemolysis. This result is not significantly different from the standard anti-inflammatory sodium diclofenac (90.09%). Oleanonic acid and ursonic acid were two similar metabolites in subfractions ethyl acetate 1, 2, and 3, which may have anti-inflammatory properties.Conclusion:R. Tomentosa leaves can have potency as an anti-inflammatory by increasing the red blood cell membrane stability equal to lysosome cells, depending on the concentration.Key Words: Anti-inflammatory, Oleanonic acid, Ursonic acid, Rhodomyrtus tomentosa     

棉花根醇提物与水提物的止咳祛痰抗炎作用研究

制备棉花根醇提物和水提物,止咳实验采用小鼠浓氨水引咳法,祛痰实验采用气管酚红排泌法,抗炎实验采用二甲苯致耳肿胀法,观察棉花根醇提物和水提物的止咳祛痰抗炎作用。实验小鼠分正常对照组,醇提物与水提物高低剂量组及阳性药物对照组。结果发现棉花根醇提物、水提物均能延长小鼠的咳嗽潜伏期,减少咳嗽次数,均能增加气管内的酚红排泌量,对二甲苯所致的耳肿胀均有显著的抑制作用。     

Antinociceptive and Anti-Inflammatory Effects of Saline Extract and Lectin-Rich Fraction from Microgramma vacciniifolia Rhizome in Mice

Previous studies have characterized a saline extract from Microgramma vacciniifolia rhizome and its lectin (MvRL)-rich fraction with low acute toxicity. In the present study, we evaluated these preparations for acute toxicity (1,000 mg/kg) and antinociceptive and anti-inflammatory activities (100–400 mg/kg for the extract and 25–50 mg/kg for the fraction). No signs of toxicity were observed. Both the extract and fraction increased the latency period for nociception in the hot plate assay, decreased writhing induced by acetic acid, and promoted analgesic effects in phases 1 and 2 of the formalin test. The antinociceptive mechanism was attributed to interactions with opioid receptors and K⁺ ATPase channels. The extract and fraction decreased carrageenan-induced paw edema in 46.15 % and 77.22 %, respectively, at the highest doses evaluated. Furthermore, the fraction was shown to act on the bradykinin pathway. The ability to decrease leukocyte migration after treatment was also verified in the peritonitis and air pouch models. In exudates collected from air pouches, decreased tumor necrosis factor (TNF)-α and increased interleukin (IL)-10 levels were noted. Both the extract and fraction also effectively inhibited the development of granulomatous tissue. In conclusion, the substances investigated in this study can be used for the development of novel therapeutic options for pain and inflammatory processes.     

Anti-Inflammatory Activity of N-Butanol Extract from Ipomoea stolonifera In Vivo and In Vitro

Ipomoea stolonifera (I. stolonifera) has been used for the treatment of inflammatory diseases including rheumatism and rheumatoid arthritis in Chinese traditional medicine. However, the anti-inflammatory activity of I. stolonifera has not been elucidated. For this reason, the anti-inflammatory activity of n-butanol extract of I. stolonifera (BE-IS) was evaluated in vivo by using acute models (croton oil-induced mouse ear edema, carrageenan-induced rat paw edema, and carrageenan-induced rat pleurisy) and chronic models (cotton pellet-induced rat granuloma, and complete Freund’s adjuvant (CFA)-induced rat arthritis). Results indicated that oral administration of BE-IS significantly attenuated croton oil-induced ear edema, decreased carrageenan-induced paw edema, reduced carrageenan-induced exudates and cellular migration, inhibited cotton pellet-induced granuloma formation and improved CFA-induced arthritis. Preliminary mechanism studies demonstrated that BE-IS decreased the levels of myeloperoxidase (MPO) and malondialdehyde (MDA), increased the activity of anti-oxidant enzyme superoxide dismutase (SOD) in vivo, and reduced the production of nitric oxide (NO), prostaglandin E₂ (PGE₂), tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-6 in lipopolysaccharide-activated RAW264.7 macrophages in vitro. Results obtained in vivo and in vitro demonstrate that BE-IS has considerable anti-inflammatory potential, which provided experimental evidences for the traditional application of Ipomoea stolonifera in inflammatory diseases.     

番石榴叶乙醇提取物中熊果酸富集的初步研究

对番石榴Psidium guajava叶乙醇提取物中熊果酸富集工艺的除杂和酸碱富集主要环节进行优化试验。结果表明,最佳富集工艺为：以料液比为1∶50的水和石油醚除去杂质;酸碱富集过程中富集条件以10% NaOH碱化提取液pH=14,再以5% H2SO4酸化调节pH=3,用1倍样品溶液体积的pH=3酸性水溶液稀释。终产品中熊果酸的纯度和回收率分别为17.74%和85.12%。     

Anti-Inflammatory Activities of Monoterpene and Sesquiterpene Glycosides from the Aqueous Extract of Artemisia annua L.

Artemisia annua L. is a Chinese medicinal herb, but the origin of its pharmacological properties, including its anti-inflammatory activity, remain unknown. In this study, five new monoterpene glycosides ( 1 – 5 ) and two new sesquiterpene glycosides ( 6 and 7 ) were isolated from the aqueous extract of the aerial parts of A. annua. The structures of these glycosides were determined using high-resolution electrospray ionization mass spectrometry, nuclear magnetic resonance spectroscopy, electronic circular dichroism calculations, and chemical hydrolysis methods. The anti-inflammatory activities of the isolated compounds were evaluated by down-regulating interleukin-6 (IL-6) in lipopolysaccharide-stimulated RAW 264.7 macrophages. Notably, all the new compounds significantly inhibited the expression of IL-6 in a dose-dependent manner.     

Anti-Inflammatory and Analgesic Activities of SKLJI,a Highly Purified and Injectable Herbal Extract of Lonicera japonica

The parenteral route has many merits over the oral route, including greater predictability, reproducibility of absorption, and rapid drug action, but injectable phytomedicines are uncommon due to protein precipitating tannin and hemolytic saponin components. In this study, in an effort to develop a safe injectable analgesic phytomedicine, we prepared a tannin and saponin-free Lonicera japonica extract, SKLJI, through fractionation and column purification, and evaluated its anti-inflammatory and analgesic activities in in vivo experimental models of inflammation and pain. The removal of tannin and saponin resulted in loganin and sweroside-enriched SKLJI and it showed reduced hemolysis and protein precipitation. In efficacy tests, SKLJI inhibited croton oil- and arachidonic acid-induced ear edema, acetic acid-induced writhing, and carrageenan-induced rat hind paw hyperalgesia. Inhibition of cylcooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), and 5-lipoxyfenase (5-LO) activities by SKLJI appeared to be the mechanism underlying anti-inflammatory and analgesic efficacy. Loganin and sweroside also showed anti-inflammatory and analgesic activities, suggesting that they might be active principles in the efficacy of SKLJI. These results suggest that SKLJI is a viable candidate for a new anti-inflammatory and analgesic phytomedicine that can be administered by the parenteral route.     

HPLC and GC Characterization of Dicliptera bupleuroides Aerial Parts and Evaluation of Its Anti-Inflammatory Potential in Vitro,in silico and in Vivo Using Carrageenan and Formalin Induced Inflammation in Rat Models

The current study aimed to evaluate the anti-inflammatory activity of Dicliptera bupleuroides Nees aerial parts methanol extract and its different fractions namely hexane, chloroform, ethyl acetate and butanol in vitro using cyclooxygenase inhibitory assay (COX-2). In vivo anti-inflammatory evaluation was performed using carrageenan and formalin induced inflammation in rat models followed by molecular docking. High performance liquid chromatography (HPLC) and gas chromatography coupled with mass chromatography (GC/MS) analyses were used for chemical analyses of the tested samples. The tested samples showed significant inhibition in COX-2 inhibitory assay where methanol extract (DBM) showed the highest inhibitory potential at 100 μg/mL estimated by 67.86 %. At a dose of 400 mg/kg, all of the examined samples showed pronounced results in carrageenan induced acute inflammation in rat model at 4^th h interval with DBM showed the highest efficiency displaying 65.32 % inhibition as compared to the untreated rats. Formalin model was employed for seven days and DBM exhibited 65.33 % and 69.39 % inhibition at 200 and 400 mg/kg, respectively approaching that of the standard on the 7^th day. HPLC revealed the presence of caffeic acid, gallic acid and sinapic acid, quercetin and myricetin in DBM. GC/MS analysis of its hexane fraction revealed the presence of 16 compounds belonging mainly to fatty acids and sterols that account for 85.26 % of the total detected compounds. Molecular docking showed that hexadecanoic acid followed by decanedioic acid and isopropyl myristate showed the best fitting within cyclooxygenase-II (COX-II) while nonacosane followed by hexatriacontane and isopropyl myristate revealed the most pronounced fitting within the 5-lipoxygenase (5-LOX) active sites. Absorption, metabolism, distribution and excretion and toxicity prediction (ADMET/ TOPKAT) concluded that most of the detected compounds showed reasonable pharmacokinetic, pharmacodynamic and toxicity properties that could be further modified to be more suitable for incorporation in pharmaceutical dosage forms combating inflammation and its undesirable consequences.     

马尾松花粉醇提物降脂作用与预防肥胖的实验研究

本实验通过观察马尾松花粉醇提物对小鼠脂质代谢的影响来探讨其抑制肥胖的初步机制.实验采用随机分组对照方法,对不同组的小鼠进行不同的干预处理.实验结果显示与高脂组(FC)组相比3个醇提物组在终体重、体重净增加和体重增加率方面都有明显降低(P＜0.01);体脂含量也有不同程度降低但未出现显著性差异;血脂中总胆固醇(TC)水平有不同程度的降低,并且甘油三酯(TG)都有明显降低(P＜0.01),高密度脂蛋白胆固醇(HDLC)水平都有显著性升高(P＜0.01);瘦素(LEP)和脂联素(ADP)水平都有不同程度升高;肝脏和脂肪组织中的肉碱棕榈酰转移酶(CPT-I)酶含量水平都得到显著升高(P＜0.01).上述结果证明马尾松花粉醇提物可以明显控制小鼠的体重增长以及改善体内脂质代谢水平,证明松花粉醇提物在抑制肥胖方面具有重要作用.     

野葛花醇提物中异黄酮含量及其抗氧化活性测定

以葛花苷为标准品,对野葛花醇提物的氯仿、乙酸乙酯、正丁醇萃取物及水溶物4部分中的异黄酮含量进行测定,并采用DPPH·法对各萃取物进行自由基清除实验.结果显示,(1)氯仿、乙酸乙酯、正丁醇萃取物及水溶物4部分的异黄酮含量分别为35.27%、53.42%、52.13%和2.34%.(2)抗氧化实验显示, 4 部分清除DPPH·的IC50值分别为252、49、197和344 μg/mL.(3)相关性分析表明,野葛花醇提物各萃取部分的抗氧化活性主要来自于其中含有的异黄酮类化合物.研究表明野葛花醇提物的乙酸乙酯和正丁醇萃取部分异黄酮含量较高;氯仿、乙酸乙酯、正丁醇萃取物及水溶物4部分对DPPH自由基均有一定的清除作用,其中以乙酸乙酯萃取物的效果最佳;表明异黄酮是一种天然的抗氧化剂,具有较强的抗氧化作用.     

Environmental Determinants of and Impact on Childhood Asthma by the Bacterial Community in Household Dust

Asthma increased dramatically in the last decades of the 20th century and is representative of chronic diseases that have been linked to altered microbial exposure and immune responses. Here we evaluate the effects of environmental exposures typically associated with asthma protection or risk on the microbial community structure of household dust (dogs, cats, and day care). PCR-denaturing gradient gel analysis (PCR-DGGE) demonstrated that the bacterial community structure in house dust is significantly impacted by the presence of dogs or cats in the home (P = 0.0190 and 0.0029, respectively) and by whether or not children attend day care (P = 0.0037). In addition, significant differences in the dust bacterial community were associated with asthma outcomes in young children, including wheezing (P = 0.0103) and specific IgE (P = 0.0184). Our findings suggest that specific bacterial populations within the community are associated with either risk or protection from asthma.Recent studies have begun to explore the microbial composition of house dust, finding great diversity and a high abundance of Gram-positive organisms (18, 24). Yet this transient community remains relatively unexplored despite increasing evidence of an association between microbial exposure and human health, in particular, the development of chronic diseases such as asthma. Settings with high levels of microbial exposure, such as farms and day care centers, have been associated with protection from asthma (3, 23), while interventions that reduce home microbial exposures have been related to higher rates of allergic sensitization (29). Moreover, variability in home microbial exposures has been linked to differences in immune response and asthma risk in childhood (1, 4). Finally, genes involved in innate immune responses to microbes have been found to interact with microbial exposures, resulting in altered risks for asthma (10). Microbial exposure has primarily been assessed in these studies through measurement of surrogates, such as lipopolysaccharide (LPS, or endotoxin), a component of the outer membrane of Gram-negative bacteria (16), and more recently N-acetyl muramic acid for Gram-positive bacteria (28) and ergosterol for fungi (25). While the use of such surrogates has confirmed a negative relationship between levels of microbial exposure and the development of asthma, exploration is needed beyond a handful of surrogates to more completely represent the complexity of the actual microbial communities present in homes.A preliminary experiment was performed to characterize the bacterial communities of dust from homes in Tucson, AZ. First, general bacterial community diversity was examined in house dust samples by using a PhyloChip microarray as described by Brodie et al. (5). These samples were obtained from homes of four volunteers in the Tucson area. Array results revealed an average of 295 ± 16 (mean ± standard deviation) unique operational taxonomic units (OTUs) per sample, where a taxon was considered present in the sample when more than 92% of the assigned probe pairs for its corresponding probe set were positive. Of these OTUs, 85% belonged to the same 13 divisions in each dust sample, with the 5 most dominant divisions including the Alphaproteobacteria (17.6 ± 2.6%), Firmicutes (16.3 ± 4.1%), Actinobacteria (13.4 ± 2.2%), Gammaproteobacteria (11.8 ± 1.1%), and Deltaproteobacteria (8.7 ± 0.4%) (see Fig. S1 in the supplemental material). Distributions among the 13 major divisions were fairly consistent but not identical among the four different dust samples. The remaining 15% of OTUs belonged to 33 other bacterial divisions but were present at frequencies of less than 1% each. Thus, the number of OTUs and the phylogenetic distributions of these OTUs among the four households were similar.A second experiment was performed to see how the structure of house dust communities varies with environmental parameters and asthma outcomes. The dust samples, examined using denaturing gradient gel electrophoresis (DGGE), were obtained from homes of infants enrolled in the Infant Immune Study (IIS), a nonselected birth cohort (12). Dust samples were collected when the child was approximately 3 months old, at which time the home condition was evaluated. Other information available on this cohort includes data on (i) early life exposures potentially related to asthma risk (the presence of a dog or cat in the home and day care attendance in the first months of life) and (ii) asthma-related outcomes of wheezing and aeroallergen sensitization (specific IgE [Sp-IgE]) (7). A subset (n = 44) of the IIS dust samples was used in this analysis. Based upon the premise that health outcomes will be influenced by the most abundant members of the dust bacterial community (in terms of numbers), DGGE of 16S rRNA gene PCR amplicons was selected as the profile technique to compare the microbial community structures of dust samples from the 44 homes. PCR-DGGE profiles evaluate 16S rRNA genes from all bacterial populations whose initial template concentrations represent >1% of the total community DNA (6, 13, 17). PhyloChip analysis was not used for this experiment because the cost would have been prohibitive for the number of households analyzed.DNA extraction was performed on the IIS dust samples by direct lysis using the Fast DNA spin kit for soil (MP Biomedicals, Solon, OH) as described previously (9). The amount of dust used for extraction ranged from 21 to 35 mg. For each set of extractions a DNA blank (no dust) was included to monitor potential contamination during the extraction process. The extracted DNA was labeled with the PicoGreen double-stranded DNA (dsDNA) quantitation reagent (Molecular Probes, Eugene, OR) and quantified using the Turner Biosystems TBS 380 fluorometer. DNA yields ranged from 0.2 to 41.2 ng DNA/mg of dust.For DGGE analysis, the community DNA obtained from the dust samples was PCR amplified (25 cycles) using the V7/V8 variable region of the 16S rRNA gene and primers 1070f and 1392R with a 40-bp GC clamp (11), following the protocol of Colores et al. (8). Unacetylated bovine serum albumin (Sigma, St. Louis, MO) was added to the PCR mixture at a concentration of 0.4 g liter⁻¹ to relieve PCR inhibition. Amplicons were separated on DGGE gels by using the Bio-Rad Laboratories system (D-Code; Hercules, CA) with a 6% acrylamide gel and a 45% to 65% urea-formamide denaturing gradient. Each DGGE gel run in this study examined either an environmental exposure or an asthma-related outcome variable, as detailed in Tables ​Tables11 and ​and2.2. For example, children reported to have actively wheezed on at least two out of three questionnaires obtained at 1, 2, and 3 years were compared with children for whom all three questionnaires were completed but for whom no wheezing was reported. Fourteen samples were analyzed on each gel, including seven different households from the negative group and seven different households from the positive group. Two lanes of the gel were also loaded with negative controls. DGGE gels were run for 15.5 h at a constant voltage of 50 V, stained with 3× SYBR green I (Molecular Probes, Eugene, OR) for 40 min, and then visualized.

TABLE 1.

Gel comparison groups for early life environmental exposures

黄兰叶乙醇提取物的活性研究和成分分析

为挖掘黄兰(Michelia champaca)叶的开发潜力,采用抗氧化评价、细胞评价和仪器分析方法对超声提取的黄兰叶乙醇提取物进行活性研究和化学成分的定性分析。DPPH·自由基清除和铁离子还原力测定结果表明,黄兰叶的20%和80%乙醇提取物具有抗氧化潜能。黄兰叶20%乙醇提取物的质量浓度超过12.5μg/mL,80%乙醇提取物的质量浓度超过6.25μg/mL,对HacaT细胞具有细胞毒性。采用超高效液相-质谱联用方法结合软件谱库搜索对黄兰叶乙醇提取物进行成分分析,共检索到206个化合物,包括黄酮、生物碱、脂肪酸、醇等多种类型,通过标准品验证可明确鉴定出芦丁、槲皮素、山奈酚等黄酮类物质。不同月份的黄兰叶乙醇提取物的成分含量不相同,成分分布在春夏季与秋冬季存在明显差异,某些成分具有季节独特性。因此,黄兰叶在日化产品开发上存在潜力,可结合实际根据成分选择采收时间。     

Effect of Sugar Concentration in Jerusalem Artichoke Extract on Kluyveromyces marxianus Growth and Ethanol Production

The effect of inulin sugars concentration on the growth and ethanol production by Kluyveromyces marxianus UCD (FST) 55-82 was studied. A maximum ethanol concentration of 102 g/liter was obtained from 250 g of sugars per liter initial concentration. The maximum specific growth rate varied from 0.44 h⁻¹ at 50 g of sugar per liter to 0.13 h⁻¹ at 300 g of sugar per liter, whereas the ethanol yield remained almost constant at 0.45 g of ethanol per g of sugars utilized.     

小鼠过敏性哮喘模型的研究进展及评价

支气管哮喘（简称哮喘）是常见的慢性病,随着过敏患者的增加,小鼠过敏性哮喘模型的研究越来越重要。本文通过对近年来国内外小鼠过敏性哮喘的实验研究文献进行总结,从实验小鼠的选择、制备模型的方法及模型的评价指标等方面进行综合分析,为进一步开展哮喘研究提供帮助。     

Differential Effects of Rapamycin and Dexamethasone in Mouse Models of Established Allergic Asthma

The mammalian target of rapamycin (mTOR) plays an important role in cell growth/differentiation, integrating environmental cues, and regulating immune responses. Our lab previously demonstrated that inhibition of mTOR with rapamycin prevented house dust mite (HDM)-induced allergic asthma in mice. Here, we utilized two treatment protocols to investigate whether rapamycin, compared to the steroid, dexamethasone, could inhibit allergic responses during the later stages of the disease process, namely allergen re-exposure and/or during progression of chronic allergic disease. In protocol 1, BALB/c mice were sensitized to HDM (three i.p. injections) and administered two intranasal HDM exposures. After 6 weeks of rest/recovery, mice were re-exposed to HDM while being treated with rapamycin or dexamethasone. In protocol 2, mice were exposed to HDM for 3 or 6 weeks and treated with rapamycin or dexamethasone during weeks 4–6. Characteristic features of allergic asthma, including IgE, goblet cells, airway hyperreactivity (AHR), inflammatory cells, cytokines/chemokines, and T cell responses were assessed. In protocol 1, both rapamycin and dexamethasone suppressed goblet cells and total CD4⁺ T cells including activated, effector, and regulatory T cells in the lung tissue, with no effect on AHR or total inflammatory cell numbers in the bronchoalveolar lavage fluid. Rapamycin also suppressed IgE, although IL-4 and eotaxin 1 levels were augmented. In protocol 2, both drugs suppressed total CD4⁺ T cells, including activated, effector, and regulatory T cells and IgE levels. IL-4, eotaxin, and inflammatory cell numbers were increased after rapamycin and no effect on AHR was observed. Dexamethasone suppressed inflammatory cell numbers, especially eosinophils, but had limited effects on AHR. We conclude that while mTOR signaling is critical during the early phases of allergic asthma, its role is much more limited once disease is established.     

Anti-Inflammatory Activity of Lauraceae Plant Species and Prediction Models Based on Their Metabolomics Profiling Data

The Lauraceae is a botanical family known for its anti-inflammatory potential. However, several species have not yet been studied. Thus, this work aimed to screen the anti-inflammatory activity of this plant family and to build statistical prediction models. The methodology was based on the statistical analysis of high-resolution liquid chromatography coupled with mass spectrometry data and the ex vivo anti-inflammatory activity of plant extracts. The ex vivo results demonstrated significant anti-inflammatory activity for several of these plants for the first time. The sample data were applied to build anti-inflammatory activity prediction models, including the partial least square acquired, artificial neural network, and stochastic gradient descent, which showed adequate fitting and predictive performance. Key anti-inflammatory markers, such as aporphine and benzylisoquinoline alkaloids were annotated with confidence level 2. Additionally, the validated prediction models proved to be useful for predicting active extracts using metabolomics data and studying their most bioactive metabolites.     

Diesel Exhaust Particles Induce Cysteine Oxidation and S-Glutathionylation in House Dust Mite Induced Murine Asthma

Background

Diesel exhaust particle (DEP) exposure enhances allergic inflammation and has been linked to the incidence of asthma. Oxidative stress on the thiol molecules cysteine (Cys) and glutathione (GSH) can promote inflammatory host responses. The effect of DEP on the thiol oxidation/reduction (redox) state in the asthmatic lung is unknown.

Objective

To determine if DEP exposure alters the Cys or GSH redox state in the asthmatic airway.

Methods

Bronchoalveolar lavage fluid was obtained from a house dust mite (HDM) induced murine asthma model exposed to DEP. GSH, glutathione disulfide (GSSG), Cys, cystine (CySS), and s-glutathionylated cysteine (CySSG) were determined by high pressure liquid chromatography.

Results

DEP co-administered with HDM, but not DEP or HDM alone, decreased total Cys, increased CySS, and increased CySSG without significantly altering GSH or GSSG.

Conclusions

DEP exposure promotes oxidation and S-glutathionylation of cysteine amino acids in the asthmatic airway, suggesting a novel mechanism by which DEP may enhance allergic inflammatory responses.