Hot water washing processing technology of Euodiae Fructus based on change laws of active components and tastes / 中国中药杂志

In order to establish the standardized processing technology of the hot water washing of Euodiae Fructus, this study, based on the traditional processing method of hot water washing of Euodiae Fructus recorded in ancient works and modern processing specifications of traditional Chinese medicine decoction pieces, took the yield of decoction pieces and the content of main components as the indicators and optimized the processing conditions by orthogonal test based on the results of single factor investigation. At the same time, electronic tongue technology was used to analyze the change law of the taste index of Euodiae Fructus during the hot water washing. The results of the single factor investigation showed that the content of the main components in Euodiae Fructus showed some regular changes during the processing. Specifically, the content of chlorogenic acid, hyperin, isorhamnetin-3-O-rutinoside, isorhamnetin-3-O-galactoside, and dehydroevodiamine decreased significantly, with average decreases of-23.75%,-27.80%,-14.04%,-14.03%, and-13.11%, respectively. The content of limonin increased significantly with an average increase of 19.83%. The content of evodiamine, rutaecarpine, evocarpine, and dihydroevocarpine showed fluctuating changes and generally increased, with average variation amplitudes of 0.54%,-3.78%, 2.69%, and 5.13%, respectively. The orthogonal test results showed that the optimum processing parameters for the hot water washing of Euodiae Fructus were as follows: washing time of 2 min, the solid-to-liquid ratio of 1∶10 g·mL~(-1), washing temperature of 80 ℃, washing once, and drying at 50 ℃. After the hot water washing processing, the average yield of Euodiae Fructus pieces was 94.80%. The content of limonin, evodiamine, and rutaecarpine was higher than those of raw pro-ducts, and the average transfer rates were 102.56%, 103.15%, and 105.16%, respectively. The content of dehydroevodiamine was lower than that of the raw products, and the average transfer rate was 83.04%. The results of taste analysis showed that the hot water washing could significantly reduce the salty, astringent, and bitter tastes of Euodiae Fructus. This study revealed the influence of the hot water washing on the content of main components and taste of Euodiae Fructus, and the processing technology of the hot water was-hing of Euodiae Fructus established in this study was stable, feasible, and suitable for industrial production, which laid a foundation for clarifying its processing principle and improving the quality standard and clinical application value of decoction pieces.

Construction of yeast cell factories for production of azadirachtin precursor tirucalla-7,24-dien-3β-ol / 中国中药杂志

Azadirachtin, as a botanical insecticide, is a highly oxidized limonoid triterpenoid existing in the seeds of Azadirachta indica. However, due to the low content in the seeds, the production of azadirachtin by seed extraction has low yield. Chemical synthesis of azadirachtin is characterized by complex process and low yield. Synthetic biology provides an alternative for the supply of azadirach-tin. In this study, two oxidosqualene cyclases AiOSC1 and MaOSC1 respectively derived from A. indica and Melia azedarach were identified in yeast. A yeast strain producing tirucalla-7,24-dien-3β-ol was constructed by integration of AiOSC1, Arabidopsis thaliana-derived squalene synthase gene(AtAQS2), and Saccharomyces cerevisiae-derived truncated 3-hydroxy-3-methyl-glutaryl coenzyme A reductase gene(PgtHMGR) into the delta site of yeast. Then, the function of MaCYP71BQ5 was successfully verified in yeast after this gene was introduced into the constructed yeast strain. This study not only laid a foundation for the biosynthesis of tirucalla-7,24-dien-3β-ol, but also provided a chassis cell for the functional identification of cytochrome oxidases(CYP450 s) in azadirachtin biosynthesis pathway.

Limonoid detection and profile in callus culture of sweet orange

Plant tissue culture has emerged as an important tool to produce bioactive compounds from various plant species, including the sustainable production of limonoids that are receiving considerable attention due to the benefits associated with human health such as anticancer activities. The purpose of the present study was to evaluate the capacity of limonoids aglycone production from callus culture from sweet orange cv. Pera (Citrus sinensis) seeds and identify the compounds produced in this cell line. Callus induction occurred in Murashige and Skoog medium supplemented with 2,4-dichlorophenoxyacetic (2,4-D), malt extract, agar and coconut water. For the analysis and identification of the limonoids, CG-MS-EI ion-positive mode and UPLC-QTOF-ESI were used operating in positive and negative mode. An intense peak corresponding to limonin appeared in the callus extracts at a retention time of 58.1 min. in CG-MS-EI and four major limonoids aglycone by positive ion mode UPLC-QTOF-ESI: limonin, nomilin, deacetylnomilin, and nomilinic acid. The culture medium was efficient at the bioproduction of limonoids aglycone in callus cultures of C. sinensis seeds. Therefore, data obtained from UPLC-QTOF-ESI proved its importance at identifying new compounds that benefit human health, and may assist future work in the identification of known or new limonoids in Citrus species and related genera.

Simultaneous Extraction and Separation of Oil and Azadirachtin from Seeds and Leaves of Azadirachta indica using Binary Solvent Extraction

Conventional extraction of oil and azadirachtin, a botanical insecticide, from Azadirachta indica involves defatting the seeds and leaves using hexane followed by azadirachtin extraction with a polar solvent. In order to simplify the process while maintaining the yield we explored a binary extraction approach using Soxhlet extraction device and hexane and ethanol as non-polar and polar solvents at various ratios and extraction times. The highest oil and azadirachtin yields were obtained at 6 h extraction time using a 50:50 solvent mixture for both neem leaves (44.7 wt%, 720 mg(Aza)/kg(leaves)) and seeds (53.5 wt%, 1045 mg(Aza)/kg(seeds)), respectively.

Two new nimbolinin- and trichilin-class limonoids isolated from the fruits of Melia azedarach / 中国天然药物

Two new furan fragment isomerized limonoids, meliazedalides A and B (compounds 1 and 2), were isolated from the fruits of Melia azedarach Linn.. Their chemical structures were elucidated on the basis of HR-ESI-MS and 1D and 2D NMR data, which belonged to nimbolinin- and trichilin-class, respectively. Compound 2 exhibited weak inhibitory effect on NO production in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages with IC being 37.41 μmol·L.

Limonoids from seeds of Azadirachta indica and their cytotoxic activity / 中国中药杂志

Eight limonoids were isolated from 95% ethanol extracts of neem(Azadirachta indica) seeds by various chromatographic methods. By comparison of their spectroscopic data with those reported in the literatures, these limonoids were determined as salannin(1), 1-detigloyl-1-isobutylsalannin(2), salannol-3-acetate(3), salannol(4), spirosendan(5), 1-detigloyloxy-3-deacetylsalannin-1-en-3-one(6), nimbin(7) and 6-deacetylnimbin(8). Compounds 2 and 5 were firstly isolated from this genus and 5 represented the only example of its type. And 6 is a new natural product. 6 showed inhibitory activity against HeLa and HL-60 cells, with IC₅₀ of(21.61±4.37) and(27.33±5.74) μmol·L⁻¹, respectively. Both 7 and 8 mildly inhibited the growth of HeLa cells, with IC₅₀ of (33.15±5.24) and (38.56±6.41) μmol·L⁻¹, respectively.

Discovery of novel limonin derivatives as potent anti-inflammatory and analgesic agents / 中国天然药物

Novel series of limonin derivatives (V-A-1-V-A-8, V-B-1-V-B-8) were synthesized by adding various tertiary amines onto the C (7)-position of limonin. The synthesized compounds possessed favorable physicochemical property, and the intrinsic solubility of the novel compounds were significantly improved, compared with limonin. Different pharmacological models were used to evaluate the analgesic and anti-inflammatory activities of the target compounds. Compound V-A-8 exhibited the strongest in vivo activity among the novel limonin analogs; its analgesic activity was more potent than aspirin and its anti-inflammatory activity was stronger than naproxen under our testing conditions.

New limonoids isolated from the bark of Melia toosendan / 中国天然药物

Two new limonoids, 12-ethoxynimbolinins G and H (compounds 1 and 2), and one known compound, toosendanin (Chuanliansu) (compound 3), were isolated from the bark of Melia toosendan. Their structures were elucidated by spectroscopic analysis and X-ray techniques. The absolute configuration of toosendanin (3) was established by single-crystal X-ray diffraction. Compounds 1-3 were evaluated for their cytotoxicity against five tumor cell lines.

Discovery of novel limonin derivatives as potent anti-inflammatory and analgesic agents / 中国天然药物

Novel series of limonin derivatives (V-A-1-V-A-8, V-B-1-V-B-8) were synthesized by adding various tertiary amines onto the C (7)-position of limonin. The synthesized compounds possessed favorable physicochemical property, and the intrinsic solubility of the novel compounds were significantly improved, compared with limonin. Different pharmacological models were used to evaluate the analgesic and anti-inflammatory activities of the target compounds. Compound V-A-8 exhibited the strongest in vivo activity among the novel limonin analogs; its analgesic activity was more potent than aspirin and its anti-inflammatory activity was stronger than naproxen under our testing conditions.

Citron Essential Oils Alleviate the Mediators Related to Rosacea Pathophysiology in Epidermal Keratinocytes

BACKGROUND: Citron is well known for an abundance of antioxidative and anti-inflammatory ingredients such as vitamin C, polyphenol compounds, flavonoids, and limonoids. OBJECTIVE: In this study, we aimed to evaluate the effects of citron essential oils on rosacea mediators in activated keratinocytes in vitro. METHODS: Normal human epidermal keratinocytes (NHEKs) were stimulated with 1α, 25-dihydroxyvitamin D3 (VD3) and interleukin 33 (IL-33) with LL-37 to induce rosacea mediators such as kallikrein 5 (KLK5), cathelicidin, vascular endothelial growth factor (VEGF), and transient receptor potential vanilloid 1 (TRPV1). These mediators were analyzed by performing reverse-transcription polymerase chain reaction (PCR), quantitative real-time PCR, immunocytofluorescence and enzyme-linked immunosorbent assay after NHEKs were treated with citron seed and unripe citron essential oils. RESULTS: The messenger RNA (mRNA) and protein levels of KLK5 and LL-37 induced by VD3 were suppressed by citron seed and unripe citron essential oils. Furthermore, the mRNA and protein levels of VEGF and TRPV1 induced by IL-33 with LL-37 were also suppressed by citron essential oils. CONCLUSION: These results show that citron essential oils have suppressive effects on rosacea mediators in activated epidermal keratinocytes, which indicates that the citron essential oils may be valuable adjuvant therapeutic agents for rosacea.

New limonoids isolated from the bark of Melia toosendan / 中国天然药物

Two new limonoids, 12-ethoxynimbolinins G and H (compounds 1 and 2), and one known compound, toosendanin (Chuanliansu) (compound 3), were isolated from the bark of Melia toosendan. Their structures were elucidated by spectroscopic analysis and X-ray techniques. The absolute configuration of toosendanin (3) was established by single-crystal X-ray diffraction. Compounds 1-3 were evaluated for their cytotoxicity against five tumor cell lines.

Effects of deoxygedunin on Alzheimer-like pathologic dysfunction induced by D-galactose combined with AlCl / 中国应用生理学杂志

OBJECTIVE@#To investigate the effects of Deoxygedunin on Aβ deposition, learning memory, and oxidative stress induced by D-galactose combined with AlCl in model rats with Alzheimer's disease and its possible mechanism.@*METHODS@#Male SD rats were randomly divided into three groups (=12):control group, model group (AD) and intervention group (AD+Deo). Morris water maze test was used to detect learning/memory and cognitive function in rats.Glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and malondialdehyde (MDA) contents in homogenate of hippocampus were detected by enzyme-linked immunosorbent assay (ELISA).Tau protein expression in rat cerebral cortex was detected by immunohistochemistry.Western blot was used to detect the expressions of extracellular signal regulated kinase 1(ERK1), protein kinase B (PKB) and tropomyosin-related kinase B (TrkB) on TrkB signaling pathway.@*RESULTS@#The results of water maze test showed that D-galactose combined with AlCl induced a significant increase in the escape latency compared with the control group (<0.05).Deoxygedunin could reverse the increase of the escape latency of the model group (<0.05).On the 7th day after removal of the platform, the model group showed an increase in escape latency compared with the control group and the intervention group (<0.01), and the number of crossing platforms was declined (<0.05); The results of immunohistochemistry and ELISA showed that the expressions of Aβ and tau protein in the model group were increased significantly compared with those of the control group (<0.01).The activities of SOD and GSH-Px were decreased significantly and the content of MDA was increased significantly.Compared with the model group, Deoxygedunin could reverse the increase of the expressions of Aβ and tau protein (<0.01), the decrease of SOD and GSH-Px activities (<0.05) and the increase of the MDA content (<0.05).Western blot results showed that Deoxygedunin treatment reversed the decreased phosphorylation levels of TrkB, AKT and ERK1 in hippocampus of the model group.@*CONCLUSIONS@#Supplement of Deoxygedunin can significantly reverse Aβ deposition, oxidative stress and cognitive deficits by activating the TrkB signal transduction pathway, which suggest that Deoxygedunin may serve as a promising therapeutic candidate for attenuating AD-like pathological dysfunction induced by D-galactose combined with AlCl.

Two new phragmalin-type limonoids orthoesters from Entandrophragma candollei / 中国天然药物

Two new phragmalin-type limonoids orthoesters, encandollens A and B (1 and 2), were isolated from the stem barks of Entandrophragma candollei collected in Ghana. The structures of these compounds were elucidated on the basis of HR-ESI-MS, H and C NMR, HSQC, HMBC, and ROESY data. Compound 1 was a rare C-15 enolic acyl phragmalin-type limonoid orthoester. Compounds 1 and 2 exhibited weak inhibitory effects on NO production in lipopolysaccharide (LPS)-induced RAW 264.7 cells.

Two new limonoids isolated from the fuits of Melia toosendan / 中国天然药物

In the present study, two new limonoids, 1α, 7α-dihydroxyl-3α-acetoxyl-12α-ethoxylnimbolinin (1) and 1α-tigloyloxy-3α-acetoxyl-7α-hydroxyl-12β-ethoxylnimbolinin (2), together with other four known limonoids (3-6), were isolated from the fruits of Melia toosendan. Their structures were elucidated by means of extensive spectroscopic analyses (NMR and ESI-MS) and comparisons with the data reported in the literature. The isolated compounds were evaluated for their antibacterial activities. Compound 4 exhibited significant antibacterial activity against an oral pathogen, Porphyromonas gingivalis ATCC 33277, with an MIC value of 15.2 μg·mL(-1). Compound 2 was also active against P. gingivalis ATCC 33277, with an MIC value of 31.25 μg·mL(-1). In conlcusion, our resutls indicate that these compounds may provide a basis for future development of novel antibiotics.

Evaluación de fenoles y limonoides en hojas de Cedrela odorata (Meliaceae) de una plantación experimental establecida en Tezonapa Veracruz, México / Evaluation of phenols and limonoids in leaves of Cedrela odorata (Meliaceae) from an experimental plantation established in Tezonapa Veracruz, Mexico

Cedrela odorata (Meliaceae) is a native timber tree to Tropical America, known for its high-quality wood, unfortunately, plantations of this species are severely attacked by Hypsipyla grandella. The attraction or repellency of this pest is related to secondary metabolites such as phenols and limonoids (triterpenes); therefore, it is important to study these compounds to understand the phytochemical phenomena behind this problem. With this aim, the concentration of total phenols and limonoides was evaluated in C. odorata leaves from a plantation established in Tezonapa Veracruz, Mexico. For this, a total of 66 tree leaves samples, from seven sites, were analyzed. Phenols and limonoids concentration showed significant differences not only among different provenances, but also among individual trees of the same site (Tukey, p≤0.05). Phenols concentration was variable and in the range from 49 to 223mg EAG/g e for total phenols, from 7 to 158mg EC/g e for flavonoids and from 4 to 104mg EC/g e for proanthocyanidins. Limonoids concentration was also variable, ranging between 227 and 748mg EL/g e. A major compound was found by High-Performance Liquid Chromatography with Ultraviolet Diode Array Detection (HPLC-UV-DAD), which corresponded to a flavonol kaempferol glycoside derivative; additionally, a flavanol catechin was also detected at low concentrations. GC-MS allowed the identification of the sesquiterpenoids β-elemene, E-caryophyllene, aromadendrene, α-humulene, γ-cadinene, D-germacrene, bicyclogermacrene, and the poly terpenoids D-α-tocopherol and β-sitosterol. Our results suggest that the evaluation of phenols may play an important role as a selection parameter for improvement and conservation programs, if they are complemented with conventional breeding practices.

Cedrela odorata (Meliaceae) es una especie forestal maderable nativa de América Tropical, conocida por la alta calidad de su madera. Plantaciones de esta especie son atacadas severamente por Hypsipyla grandella; la atracción o repelencia de la plaga está relacionada con metabolitos secundarios tipo fenoles y limonoides (triterpenos), por lo que el estudio de estos compuestos es importante para comprender algunos fenómenos fitoquímicos. Se evaluó la concentración de fenoles totales y limonoides en hojas de C. odorata (Meliaceae) de una plantación establecida en Tezonapa Veracruz México, se analizaron 66 individuos de siete procedencias. La concentración de fenoles y limonoides mostró diferencias significativas, no solo entre las procedencias sino también entre los árboles de una determinada procedencia (Tukey, p≤0.05). La concentración de fenoles totales varió de 49 a 223mg EAG/g e, los flavonoides de 7 a 158mg EC/g e y las proantocianidinas de 4 a 104mg EC/g e, mientras que en limonoides se obtuvieron valores de 227 a 748mg EL/g e. Mediante Cromatografía Líquida de Alta Resolución con detector UV-Arreglo de Diodos (HPLC-UV-DAD) se encontró un compuesto mayoritario que corresponde a un flavonol de tipo glicósido de Kaempferol y se identificó el flavanol catequina a bajas concentraciones. Por medio de Cromatografía de Gases-Espectrometría de Masas (CG-MS) se identificaron los sesquiterpenos β-elemeno, E-cariofileno, aromadendreno, humuleno, gama-cadineno, D-germacreno, biciclogermacreno y los poli terpenos Di-α-Tocoferol y β-sitosterol. Nuestros resultados sugieren que la evaluación de los fenoles puede desempeñar un papel importante como parámetro de selección en programas de mejora y conservación, si se complementan con las prácticas convencionales de mejoramiento genético.

Efeito da gedunina sobre a ativação de macrófagos induzida por LPS / Gedunin effect on the activation of macrophages induced by LPS

Os macrófagos são células do sistema imune com perfil predominantementeinflamatório, também capazes de desempenhar um papel regulatório. O lipopolissacarídeo(LPS) presente na membrana externa de bactérias Gram-negativas é um potente ativador demacrófagos, reconhecido pelo complexo de receptores membranares TLR4/MD-2/CD14, cujaativação leva a uma cascata de sinalização intracelular, dependente das proteínas adaptadorasMAL/MyD88 (via aguda) e TRAM/TRIF (via tardia). A ativação do TLR4 induz atranslocação do NFκB, a geração de diversos mediadores inflamatórios e a expressão deenzimas responsßveis pela geração de mediadores lipídicos. A gedunina, um limonóide deorigem natural presente em espécies vegetais da família Meliaceae, é um conhecidomodulador da proteína de choque térmico (Hsp)90, uma chaperona que apresenta umimportante papel na resposta imune por estabilizar diferentes proteínas clientes, tais comofatores de transcrição e tirosinas quinases, dentre as quais diversas estão associadas àsinalização do TLR4. O objetivo geral deste estudo foi avaliar o efeito da gedunina sobre aresposta inflamatória induzida por LPS in vitro e in vivo, investigando o seu papel nasinalização do TLR4 e o envolvimento da Hsp90 nesta via. O pré-tratamento in vitro demacrófagos wild type (WT) imortalizados a partir de medula óssea de camundongos C57BL/6com gedunina, em concentrações não tóxicas, inibiu o influxo de Ca2+ intracelular e aprodução de TNF-alfa e NO de forma concentração dependente, 6 h e 24 h após estímulo comLPS. Nossos dados também demonstraram que a gedunina modula tanto a via aguda quanto avia tardia da sinalização de TLR4, inibindo a produção de TNF-alfa e IL-6 em macrófagos TRIFknockout (KO) e MAL KO, 6 h e 24 h após o estímulo com LPS...

Macrophages are inflammatory cells that also play a key regulatory role in immuneresponses. Lipopolysaccharide (LPS) present on the outer membrane of Gram-negativebacteria is a potent activator of macrophages, which is recognized by the TLR4/MD-2/CD14receptor complex, triggering an intracellular signaling cascade dependent on MAL/MyD88(early phase) and TRAM/TRIF (late phase) adaptor proteins. TLR4 activation induces NFκBtranslocation into the nucleus, the production of several inflammatory mediators and theexpression of enzymes responsible for the generation of lipid mediators. Gedunin, a limonoidfrom plant species of Meliaceae family, is a modulator of the chaperone heat shock protein(Hsp)90, which in turn presents an important role in immune responses by stabilizing andactivating several eukaryotic proteins, such as transcriptional factors and tyrosine kinasesassociated with the TLR4 signaling pathway. The aim of this study was to evaluate the effectof gedunin on the inflammatory response induced by LPS in vitro and in vivo, investigating itsimpact on TLR4 cell signaling and the involvement of Hsp90 in this phenomenon. In vitropretreatment of wild type (WT) macrophages with gedunin, at non-cytotoxic concentrations,inhibited intracellular Ca2+ influx and the production of TNF-alpha and NO in a concentrationdependentmanner, 6 h and 24 h after stimulation with LPS. Our data also demonstrate thatgedunin modulates both acute and late TLR4 signaling pathways, since it inhibited TNF-alpha andIL-6 production by TRIF knockout (KO) and MAL KO macrophages, at 6 h and 24 h afterLPS stimulation...

Cytotoxicity, nitric oxide and acetylcholinesterase inhibitory activity of three limonoids isolated from Trichilia welwitschii (Meliaceae)

BACKGROUND: Limonoids are highly oxygenated compounds with a prototypical structure. Their occurrence in the plant kingdom is mainly confined to plant families of Meliaceae and Rutaceae. Owing to their wide range of pharmacological and therapeutic properties, this study was aimed at investigating the potential nitric oxide (NO) and acetylcholinesterase (AChE) inhibitory activity and the cytotoxicity of three limonoids: trichilia lactone D5 (1), rohituka 3 (2) and dregeanin DM4 (3), isolated from Trichilia welwitschii C.DC. RESULTS: Results indicated that the three limonoids had low cytotoxicity towards Vero cells with LC50 values ranging from 89.17 to 75.82 µg/mL. Compounds (2) and (3) had lower cytotoxicity compared to puromycin and doxorubicin used as reference cytotoxic compounds. Compound (1) (LC50 of 23.55 µg/mL) had good antiproliferative activity against RAW 264.7 cancer cells. At the lowest concentration tested (0.5 µg/mL), compound (2) and (3) released the lowest amount of nitric oxide (2.97 and 2.93 µM, respectively). The three limonoids had anti-AChE activity with IC50 values ranged of 19.13 µg/mL for (1), 34.15 µg/mL for (2) and 45.66 µg/mL for (3), compared to galantamine (IC50 of 8.22 µg/mL) used as positive control. CONCLUSION: The limonoid compounds studied in this work inhibited nitric oxide production in LPS-stimulated macrophages and had anti-AChE activity. Trichilia lactone D5 had potential antiproliferative activity against RAW 264.7 cancer cells. The limonoids had low cytotoxicity towards Vero cells lines. This study provided further examples of the importance of limonoids compounds as potential AChE inhibitors and anti-inflammatory agents targeting the inhibition of NO production.

Analysis on component difference in Citrus reticulata before and after being processed with salt by UPLC-Q-TOF/MS / 中国中药杂志

To analyze components of Citrus reticulata and salt-processed C. reticulata by ultra-performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF/MS), and compared the changes in components before and after being processed with salt. Principal component analysis (PCA) and partial least squares discriminant analysis (OPLS-DA) were adopted to analyze the difference in fingerprint between crude and processed C. reticulata, showing increased content of eriocitrin, limonin, nomilin and obacunone increase in salt-processed C. reticulata. Potential chemical markers were identified as limonin, obacunone and nomilin, which could be used for distinguishing index components of crude and processed C. reticulata.

Transport of limonin in rat intestine in situ and Caco-2 cells in vitro / 药学学报

Limonin existed in citrus fruits has been shown to have anti-bacterial, anti-viral, anti-feedant, anti-nociceptive, anti-inflammatory activities and anti-carcinogenic activities. But the clinical use is limited by its low bioavailability. The aim of this study is to observe the absorption and secretion transport mechanisms of limonin in intestine which can pave the way for the further study and clinical use. The transport characteristics and mechanisms of limonin in rat were studied by in situ intestine perfusion and in vitro Caco-2 cells method. The intestinal absorption of limonin was probably via a facilitated diffusion pathway which was poor and without segment-selection. Verapamil and ketoconazole improved the absorption remarkably according to the result of in vitro Caco-2 cells study; however, probenecid had no significant effect on the absorption. The P-gp efflux and CYP3A4 metabolism were involved in the poor intestinal absorption and low bioavailability of limonin. The exploration of the intestinal absorption mechanism is crucial to the design of dosage form and clinical use of limonin.

Estudo da atividade antialérgica e anti-inflamatória de tetranortriterpenóides de Carapa guianensis Aublet / Study of antiallergic activity and anti-inflammatory tetranortriterpenóides of Carapa guianensis Aublet

Carapa guianensis Aublet é um membro da família das Meliaceae amplamente utilizada na medicina popular no Brasil e em outros países que abrangem a floresta Amazônica. Nosso grupo descreveu previamente a atividade anti-inflamatória e antialérgica de um conjunto de tetranortriterpenoides (TNTPs) contendo: 6(alfa)- acetoxigedunina, 7- deacetoxi-7-oxogedunina,andiroba,anglolensato de metila e gedunina, isolados das sementes de C. guianensis. No presente estudo, foram realizados estudos in vitro e in vivo na tentativa de elucidar os mecanismos pelo quais os TNTPs aprsentam os efeitos anti-inflamatória e antialérgico, identificando assim as substâncias bioativas presentes no óleo. Nós mostramos que a incubação in vitro de esplenócitos com o conjunto de TNTPs, bem como com cada um dos TNTPS isolados foi capaz de inibir a ativação, proliferação e a produção in vitro de IL-2, CCL11/Eotaxina e CCL5/RANTES em linfócitos T através do bloqueio da translocação do fator NfkB. Além disso, a incubação in vitro de eosinófilos com o conjunto de TNTPs, bem como com cada um dos cinco TNTPs isolados,inibiu a adesão e quimiotaxia destas células. Consistente com estes dados, nós demonstramos que a gedunina apresenta uma importante atividade antialérgica em modelos in vitro de inflamação alérgica ( pleurisia alérgica e inflamação aguda pulmonar alérgica). Descobrimos que a pleurisia alégica induzida por ovalbumina (OVA) em camundongos pré- sensibilizados foi significativamente reduzida com o pré-tratamento com gedunina; esta inibição foi principalmente devido ao bloqueio do influxo de eosinófilos e de linfócitos T. O pré-tratamento com gedunina diminuiu os níveis de CCL2,CCL3,CCL5,CCL11,IL-5 e LTB4 na cavidade pleural. O pré-tratamento in vitro com gedunina reduziu o número de linfócitos T na pleura e no linfonodo torácico após 24h do desafio com OVA. O tratamento in vitro de linfócitos T com gedunina inibiu a ativação,proliferação,produção de IL2 e translocação de NfkB e de NFAT induzido por (alfa)-CD3 mAb. Notavelmente o pós-tratamento com gedunina reverteu a inflamação aguda pulmonar alérgica induzida por OVA, diminuindo as contagens de linfócitos T e eosinófilos; e níveisde mediadores eosinofilotáticos no lavabo broncoalveolar. Nosso grupo previamente demonstrou que a endotelina(ET)-1 desempenha importante papel pró-inflamatório na resposta inflamatória articular aguda. Desta forma, nós avaliaremos os efeitos anti-inflamatório e -nociceptivos da gedunina na artrite induzida por zymosan e seu efeito biológico na expressão de ET-1. A injeção intra-articular com zymosan induz a expressão de RNAm para prepoET-1, levando a produção de TNF(alfa),LTB4 e IL-6 resultando no influxo de neutrófilos, formação de edema e hiperalgesia. Pré e pós-tratamento com gedunina diminuiram o acúmulo de neutrófilos, inchaço na articulação do joelho, expressão de RNAm para preproET-1 e produção de mediadores inflamatórios. Além diso, o pré-tratamento in vitor com gedunina inibiu a ativação e quimiotaxia de neutrófilos induzida por ET-1. Em conclusão, o presente estudo mostrou que os TNTPs apresentam notável atividade anti-inflamatória e antialérgica inibindo a ativação e migração de leucócitos; e bloqueando deferentes mediadores pró-inflamatórios.