Citrus aurantium for inflammation -
aurantifolia RFA , C. limon RFA and C. latifolia RFA The fruit peels were manually removed and homogenized with water in a blender, then immediately submitted to hydro distillation in a modified Clevenger-type apparatus for 2 hours.
After extraction, the oils were dried over anhydrous Na 2 SO 4 and stored at °C. Typically, g of fruit peels were used and average yields of 1. GC analyses were carried out using a Shimadzu GC Tokyo, Japan equipped with a flame ionization detector on a DB5 fused silica capillary column 30 m, 0.
Injector and detector temperatures were maintained at °C and °C, respectively. The oil samples were dissolved in CHCl 3 , and 1 μL aliquots were injected in split mode with split ratio of using H 2 as the carrier gas 1.
Kovats retention indices KI of the compounds were determined relative to the retention times of a series of n-alkanes C7—C30 with linear interpolation. The relative amounts of the components were calculated based on GC peak areas without correction factors. Analyses were made on a Shimadzu QP Plus system using a DB5-MS fused silica capillary column 30 m, 0.
The carrier gas was He Oil samples were diluted in CHCl 3 and 1 μL aliquots were injected in split mode split ratio The identification of individual components was based on i a comparison of the mass spectral fragmentation patterns with those stored in the NIST Mass Spectral Library and ii a comparison of the GC Kovats retention indices KI on a DB-5 column with those of authentic compounds and from literature data [ 13 ].
The characterization of the isolated compounds was made by comparison of their 13 C NMR spectra CDCl 3 , MHz with literature data [ 14 ].
Male Swiss Webster mice 2 months old, 18—25 g , kindly donated by Instituto Vital Brazil, were used in this study. Twelve hours before each experiment, the animals received only water in order to avoid food interference with substances absorption.
The animals were acclimatized to the laboratory for at least 1 h before testing and were used only once throughout the experiments. Physical condition of animals was daily monitored and animals with any signs of suffering were euthanized.
Also none of animals used became severely ill or died at any time prior to the experimental endpoint. The research was conducted in accordance with the internationally accepted principles for laboratory animal use and care.
All solvents were with chromatographic grade Tedia, Brazil. Acetylsalicylic acid ASA , carrageenan, dexamethasone, citral and limonene were purchased from Sigma-Aldrich St. Louis, MO, U. Formalin was purchased from Merck Germany. Morphine sulfate was kindly provided by Cristália São Paulo, Brazil.
and dexamethasone 1. were used as reference drugs. All drugs were diluted in phosphate buffer saline PBS just before use. The control group was composed by vehicle PBS with the same amount of oil used in the highest dose. The final concentration of oil did not exceed 0. Mice were tested according to the method described by Sahley and Berntson [ 15 ] 5 and adapted by Matheus et al.
Animals were placed on a hot plate Insight Equipment, Brazil set at 55 ± 1°C. At successive intervals of 30 min after oral administration of EOs or vehicle, the reaction time was recorded when the animals licked their fore- and hind-paws and jumped.
Baseline was considered the mean reaction time obtained at 60 and 30 min before administration of the compounds, vehicle, or morphine and was defined as the normal reaction of the animal to the temperature.
When animals were kept on the hot plate for a period of time greater than three times the baseline cut-off , they were removed to avoid possible damage to the paws. The licking behavior was examined immediately after injection of formalin into the hind paw.
The procedure was similar to the method described by Hunskaar and Hole [ 17 ] and adapted by Gomes et al. Mice received an injection of 20 μL of formalin 2. The time that the animal spent licking the injected paw was immediately recorded. The nociceptive and inflammatory response consists of the following two phases: the first phase lasts until 5 min after the formalin injection first phase, neurogenic pain response , and the second phase occurs 15—30 min after the formalin injection second phase, inflammatory pain response.
The animals were pre-treated with oral doses of EOs, vehicle or ASA for 60 min before the administration of formalin. Animals were used as described by Sedgwick et al. treated groups 1 h before carrageenan injection. Another vehicle-treated group was used in mice that received PBS phosphate buffer saline, 1 mL in SAP.
After 24 h all groups were sacrificed, SAP was washed with 1 mL of sterile PBS and exudates collected. Bone marrow cells were obtained by flushing the femoral cavity with 1 mL of phosphate buffer saline PBS.
Peripheral blood was collected in a heparinized tube. Cells counts in from aliquot of bone marrow cells, blood cells suspension or exudates were determined in a CellPocHiV Diff Sysmex haematology analyser. The exudates were also centrifuged 5, x g , 10 min, 4°C and aliquots of the supernatants were stored at °C until the assays.
To evaluate the production of nitric oxide NO , the nitrate accumulated in the SAP exudates was measured according to the method described by Bartholomew [ 21 ] and adapted by Raymundo et al [ 20 ]. FlexStation microplate reader Molecular Devices, USA was used to measure the absorbance at nm.
Nitrite concentration was measured by comparison with a standard curve of sodium nitrite. To evaluate a possible toxic effect we adapted the method used by Lorke [ 23 ].
During 5 consecutive days several parameters i. The stomachs of the animals were removed and opened to observe any signs of hyperemia and the presence or absence of ulcer. All experimental groups consisted of 6—10 mice. The results are presented as the mean ± S.
P values less than 0. Table 1 show retention indices and percentages of each compound identified in the essential oils of the Citrus species studied. Twenty-one to thirty-one compounds were identified, comprising Limonene The sesquiterpenes β-bisabolene, trans-α-bergamotene and β-caryophyllene were present in the essential oils of all the citrus studied in low amounts.
The majority of compounds identified are hydrocarbon monoterpenes. Kovats indexes KI were compared with literature [ 25 ]. In order to evaluate possible anti-inflammatory or antinociceptive effects of the EOs obtained from C. latifolia and C. limonia , the first model used was the formalin-induced paw licking.
The first phase developed during the first 5 minutes after formalin injection, with mice in the control group continuing to lick the paw for The second phase developed between 15 and 30 minutes after formalin injection, with mice continuing to lick the injected paw for aurantifolia EOs Fig 1.
To rule out a possible central antinociceptive activity from the EOs, we also evaluated their effects in the hot plate model. Statistical significance was calculated by ANOVA followed by Bonferroni's test. Based on the results of the formalin-induced licking behavior, we decided to further test smaller doses of those EOs that demonstrated a significant effect i.
The positive control group used acetylsalicylic acid, ASA significantly reduced licking-response at the 2 nd phase. Next, we analyzed the capacity of the EOs to reduce cell migration into the subcutaneous air pouch SAP after the injection of carrageenan. The results obtained in this model show that C.
Fig 3. Reduction in cells number inhibited both mononuclear and polymorphonuclear leukocytes without distinction between then data not shown. Because both EOs significantly reduced cell migration into the SAP, we decided to further analyze other parameters present in the inflammatory processes induced by carrageenan.
We therefore measured the amount of nitric oxide NO produced and the amount of protein extravasated to the exudate in the cavity. Both C.
aurantifolia EOs significantly reduced the amount of protein extravasated and NO produced in all three doses evaluated with results similar to those obtained after pretreatment of animals with dexamethasone Fig 4.
or vehicle. Fig 5 shows the results of the cytokine quantification in the SAP exudate. Carrageenan injection in the SAP induced limon significantly reduced the amount of IL-1β in the SAP, and all three doses inhibited TNF-α and IFN-γ production.
Levels of IL-1β in the exsudates were reduced with pretreatment of mice with higher dose of C. Pretreatment of mice with dexamethasone also significantly reduce cytokine levels. As the EOs significantly reduced cell migration into the SAP, we decided to evaluate a possible cytotoxic effect against blood and bone marrow leukocytes.
limon or C. However, a significant reduction in leukocytes counts in blood and bone marrow were observed after treatment with the same dose of C. aurantifolia EO, suggesting cell toxicity Fig 6. The unexpected effect observed in C. aurantifolia led us to investigate the possible causes.
As previously shown, all three EOs contain citral. aurantifolia EO contain 9. limon EO contains 2. These doses corresponded to the amount obtained from C. aurantifolia EO. As can be observed in Fig 6 , all three doses of citral significantly reduced the number of leukocytes in blood and bone marrow.
Due to the cytotoxic effect against the blood and bone marrow cells we decided to evaluate the possible toxicity in behavioral parameters in mice.
After finding high amounts of limonene in the EOs limon , limonia and aurantifolia , we decided to evaluate pure limonene.
The doses used were similar to the amount found in C. In the SAP model, TNF-α levels were inhibited by all three doses 5. None of the doses tested influenced the total leukocyte counts in blood or bone marrow data not shown.
Animals were pretreated with various doses of limonene 5. In this study we demonstrated that essential oils from several citrus species presented significant anti-inflammatory effects in different models in mice.
To confirm the hypothesis that essential oil EO from C. limonia have antinociceptive effects, we tested each one on the formalin-induced licking behavior. This model is a widely-used pain model in evaluating antinociceptive and anti-inflammatory drugs.
There are two phases, with the first neurogenic phase occurring peripherally and resulting from formalin activation of nociceptors located in the tissue and the second inflammatory phase occurring after the release of a multitude of molecules, such as histamine, serotonin, and bradykinin, developing an inflammatory response [ 25 ].
Our results suggest that Eos from C. limonia have an anti-inflammatory effect because they reduced the second phase response to formalin. This may occur through a reduction in inflammatory mediator liberation in mice paws or a direct action on one or more mediator receptors.
The formalin model alone cannot be used to ascertain the anti-inflammatory effects of the Citrus species. We therefore used the carrageenan-induced inflammation in a subcutaneous air pouch SAP model. This model is characterized by a drastic increase in leukocytes, cytokines, inflammatory mediators and protein after the carrageenan injection [ 20 ].
The mechanism by which carrageenan induces the inflammatory response is complex and involves the liberation of several mediators and an increase in vascular permeability. Our results indicate that EOs induce an anti-inflammatory effect by reducing several of the parameters observed in carrageenan-induced inflammation.
These results cannot guarantee the exact local action of the EOs, but do suggest that EOs can be acting in one or more mediator systems involved in the inflammation. During the inflammatory event there is also an increase in cytokine production.
Tumor necrosis factor-α TNF-α , interleukin-1β IL-1β , and interferon-γ IFN-γ have important roles in the maintenance of the inflammatory profile [ 26 , 27 ]. Our results indicate that EOs have significant anti-inflammatory effects in reducing all parameters evaluated.
Since IL-1β and TNF-α regulate leukocyte migration, we can infer that a reduction in leukocyte number may have been a direct effect of EOs as well as an indirect effect resulting from the reduction in the levels of those cytokines.
Taken together, these results indicate that these EOs act similarly to immunomodulators in reducing cell migration and inflammatory mediator production. Our results are in agreement with others that showed various essential oils or their constituents inhibit cytokine production. For example, 1,8-cineol inhibited TNF-α and IL-1β in human lymphocytes, α-humulene reduced TNF-α production, and terpinenol suppressed the production of TNF-α, IL-1β, IL-8, IL, and PGE2 by LPS-activated monocytes [ 28 ].
latifolia reduced TNF-α and IL levels in zymosan-induced peritonitis [ 9 ]. Despite all EOs present limonene in there constitution a difference occurred between the inhibitions observed in the formalin-induced licking and carrageenan-induced cell migration models.
A possible explanation to the differences could be the fact that both models are very different. In the first one we observe liberation of inflammatory mediators such as histamine, serotonin and bradykinin in mice paws [ 17 ]. In the second model we have the involvement of several other mediators and systems, liberation of nitrogen reactive species ROS and cytokines [ 19 ].
Additionally, C. aurantifolia EO demonstrated some toxicity, likely due to the presence of high amounts of citral. Others have demonstrated a toxic effect of citral. However, those groups used different models such as embryofetal [ 29 ] and carcinogenesis induction [ 30 ]. Although very different models and doses were used, our data, in our experimental conditions, indicate that citral has toxic effect in other model and indicate that this effect is not a model-specific effect.
With our acute assays it is not possible to definitively conclude that there is a myelotoxicity given that such effects are usually studied in repeated dose toxicity studies, as e.
given in the FDA guideline. It seems premature to conclude definitively that there is myelotoxicity also in other conditions. The observed phenomena could also be a temporary reaction which would resume later, so that further studies in models for repeated dose toxicity and e. also in another animal species seem adequate.
In this regard, further studies are necessary before drawing so far reaching conclusions, given the wide spread occurrence of citral in many plants as in food as in medicinal use. We further showed that C. limonia have significant effects, likely due to the presence of high amounts of limonene The fact that EO from C.
limonia presented significant anti-inflammatory effect probably due to the presence of limonene latifolia did not present this effect even with Or it could be that any other substance present in a different amount than in others EO could interfere with an effect.
Several pharmacological effects for limonene have been documented, such as in vitro inhibition of NO and PGE2 production [ 31 ], antineoplastic [ 32 ] and anti-inflammatory in a colitis model [ 33 ], but to the best of our knowledge this work is the first to describe its effect as an inhibitor of cell migration and cytokine production.
Our results indicate that the essential oils obtained from Citrus limon , Citrus limonia and Citrus aurantifolia demonstrate a significant anti-inflammatory effect. We also draw attention to the fact that C. aurantifolia is rich in citral, which gives it a toxic and myelotoxic effects. In this regard, care should be taken with C.
aurantifolia due to its potential toxic effect. The essential oils compositions found for the species included in this report are inside the range of compositions already published for those species [ 3 ]. Additionally all the species are cultivated, meaning that we have homogenous genetic matrices as oil sources.
However, due to the well-recognized variability in the composition of secondary metabolites caused by the susceptibility of plants to, for example, seasonal, geographical and geographical influence the potential applicability of these oils to treat inflammatory conditions would gain efficacy and safety after the standardization of production of these oils focusing on the best composition for therapeutic applications.
Animals were orally pretreated with different doses of each essential oil or vehicle. The results are presented as mean ± S. We would like to thank Mr. Alan Minho for technical assistance, Dr.
Graciela Donald for English revision and Instituto Vital Brazil Niteroi, Brazil for donation of mice. Conceived and designed the experiments: AJRS PDF.
Performed the experiments: JLA DLRS MMGP DSAM CSA. Analyzed the data: JLA AJRS PDF. Wrote the paper: CSA DSAM AJRS PDF. Browse Subject Areas? Click through the PLOS taxonomy to find articles in your field.
Article Authors Metrics Comments Media Coverage Reader Comments Figures. Abstract Citrus fruits have potential health-promoting properties and their essential oils have long been used in several applications.
Introduction As rich sources of dietary fiber, vitamin C, phenols, and flavonoids, Citrus fruits are believed to have potential health-promoting properties [ 1 ]. Methods Plant material C. The oil also significantly reduced carrageenan-induced paw edema in rats.
In cotton pellet-induced granuloma, neroli was effective regarding the transudate and granuloma formation amount.
Neroli was analyzed by gas chromatography GC and gas chromatography-mass spectrometry GC-MS and twenty-three constituents, representing The major components of neroli were characterized as linalool The results suggest that neroli possesses biologically active constituent s that have significant activity against acute and especially chronic inflammation, and have central and peripheral antinociceptive effects which support the ethnomedicinal claims of the use of the plant in the management of pain and inflammation.
Abstract The analgesic and anti-inflammatory properties of Citrus aurantium L. Publication types Research Support, Non-U.
Although traditionally Cittus to Energy enhancing products indigestion, diarrhea, aurantjum, and Thermogenic weight loss capsules, the Energy enhancing products effects of Citrus aurantium on intestinal inflammation Cittus Energy enhancing products. Inflammaion aim of this study was to evaluate the aurantiuj effects and to identify the active components of a hydroalcoholic extract of C. aurantium HECA on ulcerative colitis. Animals were randomly assigned to one of four groups based on the treatment conditions. Ulcerative colitis was induced by administration of 2. Body weight, clinical signs, colon length, pro-inflammatory cytokine expression levels, and histopathological findings were evaluated. HECA markedly protected against body weight loss and colon shortening.Essential oil has been popularly used as an airantium for foor treatment of Citrys. Energy enhancing products bioactivities of essential Ctrus Citrus aurantium for inflammation blossoms of Citrus aurantium for inflammation aurantium L.
inflammatiion Engl CAVAO showed greater anti-inflammation indlammation than Detoxification Retreats Worldwide of antioxidant, anticancer, and Energy enhancing products foe inhibition.
CAVAO also markedly decreased the expression levels of cyclooxygenase-2 COX-2 Citrus aurantium for inflammation and Energy enhancing products. Furthermore, CAVAO inhibited nuclear factor-κB CCitrus activation, which was justified by the inhibitory effect on NF-κB nuclear translocation, IκBα phosphorylation and degradation, and phosphorylation-dependent IκB kinase activation in RAW CAVAO also suppressed the phosphorylation of c-Jun N-terminal kinase JNK and p38, indicating that mitogen-activated protein kinase MAPK signaling pathways were also blocked.
The major constituents of CAVAO were characterized as linalool It is concluded that CAVAO has great potential to be developed into a functional food for the treatment of inflammatory-associated diseases. Keywords: MAPK; NF-κB; RAW Abstract Essential oil has been popularly used as an alternative for the treatment of inflammation.
Substances Acyclic Monoterpenes Anti-Inflammatory Agents Cyclohexane Monoterpenes Cyclohexenes Interleukin-1beta Interleukin-6 Lipopolysaccharides Monoterpenes NF-kappa B Oils, Volatile Terpenes Tumor Necrosis Factor-alpha alpha-terpineol Nitric Oxide linalyl acetate Limonene linalool Nitric Oxide Synthase Type II Cyclooxygenase 2.
: Citrus aurantium for inflammation| Anti-inflammatory Effect of Essential Oil from Citrus aurantium L. var. amara Engl | Mar 17, Written By Amber Charles Alexis, Auranyium, RDN. Citrus aurantium for inflammation Immunol ; 16 Khan Aurantiun, Motomura Citrus aurantium for inflammation, Wang H, El-Sharkawy RT, Antioxidant-rich skincare EF, Verma-Gandhu M, Rollins BJ, Inflammationn SM. Auraantium Energy enhancing products of the review are uploaded in TIF format with dpi resolution and RGB color mode. The sesquiterpenes β-bisabolene, trans-α-bergamotene and β-caryophyllene were present in the essential oils of all the citrus studied in low amounts. They work by deactivating free radicals, which are unstable compounds that damage your cells, increasing inflammation and your disease risk 15 As mentioned above, CACH contains abundant flavonoids, including neohesperidin 4 and PMFs Keywords: MAPK; NF-κB; RAW |
| What Is Bitter Orange, and Does It Aid Weight Loss? | blossoms essential oil neroli were investigated in mice and rats. The analgesic activity of neroli was assessed by acetic acid-induced writhing and Eddy's hot plate methods, while acute and chronic anti-inflammatory effects were investigated by inflammatory paw edema in rat and the cotton pellet-induced granuloma tissue model, respectively. Mechanistic studies were conducted using L-nitro arginine methyl ester L-NAME , an inhibitor of NO synthase. Neroli significantly decreased the number of acetic acid-induced writhes in mice compared to animals that received vehicle only. Also, it exhibited a central analgesic effect, as evidenced by a significant increase in reaction time in the hot plate method. The oil also significantly reduced carrageenan-induced paw edema in rats. In cotton pellet-induced granuloma, neroli was effective regarding the transudate and granuloma formation amount. If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given. If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. Read more about how to correctly acknowledge RSC content. Fetching data from CrossRef. This may take some time to load. Loading related content. Jump to main content. Jump to site search. You do not have JavaScript enabled. Please enable JavaScript to access the full features of the site or access our non-JavaScript page. Issue 6, Bergaptol from blossoms of Citrus aurantium L. You have access to this article. Please wait while we load your content Something went wrong. Try again? Cited by. Download options Please wait Supplementary information PDF K. Article type Paper. Submitted 07 Feb Accepted 28 Apr First published 30 Apr Download Citation. Food Funct. Request permissions. |
| REVIEW article | Neroli significantly decreased the number of acetic acid-induced writhes in mice compared to animals that received vehicle only. Also, it exhibited a central analgesic effect, as evidenced by a significant increase in reaction time in the hot plate method. The oil also significantly reduced carrageenan-induced paw edema in rats. In cotton pellet-induced granuloma, neroli was effective regarding the transudate and granuloma formation amount. Monocyte chemoattractant protein-1 contributes to gut homeostasis and intestinal inflammation by composition of ILproducing regulatory macrophage subset. J Immunol ; 16 Khan WI, Motomura Y, Wang H, El-Sharkawy RT, Verdu EF, Verma-Gandhu M, Rollins BJ, Collins SM. Critical role of MCP-1 in the pathogenesis of experimental colitis in the context of immune and enterochromaffin cells. Am J Physiol Gastrointest Liver Physiol ; GG 17 Zhang Z, Yang L, Wang B, Zhang L, Zhang Q, Li D, Zhang S, Gao H, Wang X. Protective role of liriodendrin in mice with dextran sulphate sodium-induced ulcerative colitis. Int Immunopharmacol ; 18 Brito A, Ramirez JE, Areche C, Sepulveda B, Simirgiotis MJ. HPLC-UV-MS profiles of phenolic compounds and antioxidant activity of fruits from three citrus species consumed in Northern Chile. Molecules ; 19 Guccione C, Bergonzi MC, Piazzini V, Bilia AR. A simple and rapid HPLC-PDA MS method for the profiling of Citrus peels and traditional Italian liquors. Planta Med ; 20 Nelson BC, Putzbach K, Sharpless KE, Sander LC. Mass spectrometric determination of the predominant adrenergic protoalkaloids in bitter orange Citrus aurantium. J Agric Food Chem ; 21 Jerkovic I, Druzic J, Marijanovic Z, Gugic M, Jokic S, Roje M. sinensis cv. Washington navel, C. Tarocco and C. Doppio Sanguigno from Dubrovnik Area Croatia. Nat Prod Commun ; 22 Russo M, Torre G, Carnovale C, Bonaccorsi I, Mondello L, Dugo P. A new HPLC method developed for the analysis of oxygen heterocyclic compounds in Citrus essential oils. J Essent Oil Res ; 23 Arbona V, Iglesias DJ, Gomez-Cadenas A. Non-targeted metabolite profiling of Citrus juices as a tool for variety discrimination and metabolite flow analysis. BMC Plant Biol ; 38 24 Al-Rejaie SS, Abuohashish HM, Al-Enazi MM, Al-Assaf AH, Parmar MY, Ahmed MM. Protective effect of naringenin on acetic acid-induced ulcerative colitis in rats. World J Gastroenterol ; 25 Eun SH, Woo JT, Kim DH. Tangeretin inhibits IL expression and NF- κ B activation in dendritic cells and attenuates colitis in mice. Planta Med ; 26 Habtemariam S, Belai A. Natural therapies of the inflammatory bowel disease: the case of rutin and its aglycone, quercetin. Mini Rev Med Chem ; 27 Ju S, Ge Y, Li P, Tian X, Wang H, Zheng X, Ju S. Dietary quercetin ameliorates experimental colitis in mouse by remodeling the function of colonic macrophages via a heme oxygenasedependent pathway. Cell Cycle ; 4: 28 Kang GD, Kim DH. J Ethnopharmacol ; 29 Kangawa Y, Yoshida T, Abe H, Seto Y, Miyashita T, Nakamura M, Kihara T, Hayashi SM, Shibutani M. Anti-inflammatory effects of the selective phosphodiesterase 3 inhibitor, cilostazol, and antioxidants, enzymatically-modified isoquercitrin and α -lipoic acid, reduce dextran sulphate sodium-induced colorectal mucosal injury in mice. Exp Toxicol Pathol ; 30 Kumar VS, Rajmane AR, Adil M, Kandhare AD, Ghosh P, Bodhankar SL. Naringin ameliorates acetic acid induced colitis through modulation of endogenous oxido-nitrosative balance and DNA damage in rats. J Biomed Res ; 31 Marquez-Flores YK, Villegas I, Cardeno A, Rosillo MA, Alarcon-de-la-Lastra C. Acetylsalicylic acid ASA , carrageenan, dexamethasone, citral and limonene were purchased from Sigma-Aldrich St. Louis, MO, U. Formalin was purchased from Merck Germany. Morphine sulfate was kindly provided by Cristália São Paulo, Brazil. and dexamethasone 1. were used as reference drugs. All drugs were diluted in phosphate buffer saline PBS just before use. The control group was composed by vehicle PBS with the same amount of oil used in the highest dose. The final concentration of oil did not exceed 0. Mice were tested according to the method described by Sahley and Berntson [ 15 ] 5 and adapted by Matheus et al. Animals were placed on a hot plate Insight Equipment, Brazil set at 55 ± 1°C. At successive intervals of 30 min after oral administration of EOs or vehicle, the reaction time was recorded when the animals licked their fore- and hind-paws and jumped. Baseline was considered the mean reaction time obtained at 60 and 30 min before administration of the compounds, vehicle, or morphine and was defined as the normal reaction of the animal to the temperature. When animals were kept on the hot plate for a period of time greater than three times the baseline cut-off , they were removed to avoid possible damage to the paws. The licking behavior was examined immediately after injection of formalin into the hind paw. The procedure was similar to the method described by Hunskaar and Hole [ 17 ] and adapted by Gomes et al. Mice received an injection of 20 μL of formalin 2. The time that the animal spent licking the injected paw was immediately recorded. The nociceptive and inflammatory response consists of the following two phases: the first phase lasts until 5 min after the formalin injection first phase, neurogenic pain response , and the second phase occurs 15—30 min after the formalin injection second phase, inflammatory pain response. The animals were pre-treated with oral doses of EOs, vehicle or ASA for 60 min before the administration of formalin. Animals were used as described by Sedgwick et al. treated groups 1 h before carrageenan injection. Another vehicle-treated group was used in mice that received PBS phosphate buffer saline, 1 mL in SAP. After 24 h all groups were sacrificed, SAP was washed with 1 mL of sterile PBS and exudates collected. Bone marrow cells were obtained by flushing the femoral cavity with 1 mL of phosphate buffer saline PBS. Peripheral blood was collected in a heparinized tube. Cells counts in from aliquot of bone marrow cells, blood cells suspension or exudates were determined in a CellPocHiV Diff Sysmex haematology analyser. The exudates were also centrifuged 5, x g , 10 min, 4°C and aliquots of the supernatants were stored at °C until the assays. To evaluate the production of nitric oxide NO , the nitrate accumulated in the SAP exudates was measured according to the method described by Bartholomew [ 21 ] and adapted by Raymundo et al [ 20 ]. FlexStation microplate reader Molecular Devices, USA was used to measure the absorbance at nm. Nitrite concentration was measured by comparison with a standard curve of sodium nitrite. To evaluate a possible toxic effect we adapted the method used by Lorke [ 23 ]. During 5 consecutive days several parameters i. The stomachs of the animals were removed and opened to observe any signs of hyperemia and the presence or absence of ulcer. All experimental groups consisted of 6—10 mice. The results are presented as the mean ± S. P values less than 0. Table 1 show retention indices and percentages of each compound identified in the essential oils of the Citrus species studied. Twenty-one to thirty-one compounds were identified, comprising Limonene The sesquiterpenes β-bisabolene, trans-α-bergamotene and β-caryophyllene were present in the essential oils of all the citrus studied in low amounts. The majority of compounds identified are hydrocarbon monoterpenes. Kovats indexes KI were compared with literature [ 25 ]. In order to evaluate possible anti-inflammatory or antinociceptive effects of the EOs obtained from C. latifolia and C. limonia , the first model used was the formalin-induced paw licking. The first phase developed during the first 5 minutes after formalin injection, with mice in the control group continuing to lick the paw for The second phase developed between 15 and 30 minutes after formalin injection, with mice continuing to lick the injected paw for aurantifolia EOs Fig 1. To rule out a possible central antinociceptive activity from the EOs, we also evaluated their effects in the hot plate model. Statistical significance was calculated by ANOVA followed by Bonferroni's test. Based on the results of the formalin-induced licking behavior, we decided to further test smaller doses of those EOs that demonstrated a significant effect i. The positive control group used acetylsalicylic acid, ASA significantly reduced licking-response at the 2 nd phase. Next, we analyzed the capacity of the EOs to reduce cell migration into the subcutaneous air pouch SAP after the injection of carrageenan. The results obtained in this model show that C. Fig 3. Reduction in cells number inhibited both mononuclear and polymorphonuclear leukocytes without distinction between then data not shown. Because both EOs significantly reduced cell migration into the SAP, we decided to further analyze other parameters present in the inflammatory processes induced by carrageenan. We therefore measured the amount of nitric oxide NO produced and the amount of protein extravasated to the exudate in the cavity. Both C. aurantifolia EOs significantly reduced the amount of protein extravasated and NO produced in all three doses evaluated with results similar to those obtained after pretreatment of animals with dexamethasone Fig 4. or vehicle. Fig 5 shows the results of the cytokine quantification in the SAP exudate. Carrageenan injection in the SAP induced limon significantly reduced the amount of IL-1β in the SAP, and all three doses inhibited TNF-α and IFN-γ production. Levels of IL-1β in the exsudates were reduced with pretreatment of mice with higher dose of C. Pretreatment of mice with dexamethasone also significantly reduce cytokine levels. As the EOs significantly reduced cell migration into the SAP, we decided to evaluate a possible cytotoxic effect against blood and bone marrow leukocytes. limon or C. However, a significant reduction in leukocytes counts in blood and bone marrow were observed after treatment with the same dose of C. aurantifolia EO, suggesting cell toxicity Fig 6. The unexpected effect observed in C. aurantifolia led us to investigate the possible causes. As previously shown, all three EOs contain citral. aurantifolia EO contain 9. limon EO contains 2. These doses corresponded to the amount obtained from C. aurantifolia EO. As can be observed in Fig 6 , all three doses of citral significantly reduced the number of leukocytes in blood and bone marrow. Due to the cytotoxic effect against the blood and bone marrow cells we decided to evaluate the possible toxicity in behavioral parameters in mice. After finding high amounts of limonene in the EOs limon , limonia and aurantifolia , we decided to evaluate pure limonene. The doses used were similar to the amount found in C. In the SAP model, TNF-α levels were inhibited by all three doses 5. None of the doses tested influenced the total leukocyte counts in blood or bone marrow data not shown. Animals were pretreated with various doses of limonene 5. In this study we demonstrated that essential oils from several citrus species presented significant anti-inflammatory effects in different models in mice. To confirm the hypothesis that essential oil EO from C. limonia have antinociceptive effects, we tested each one on the formalin-induced licking behavior. This model is a widely-used pain model in evaluating antinociceptive and anti-inflammatory drugs. There are two phases, with the first neurogenic phase occurring peripherally and resulting from formalin activation of nociceptors located in the tissue and the second inflammatory phase occurring after the release of a multitude of molecules, such as histamine, serotonin, and bradykinin, developing an inflammatory response [ 25 ]. Our results suggest that Eos from C. limonia have an anti-inflammatory effect because they reduced the second phase response to formalin. This may occur through a reduction in inflammatory mediator liberation in mice paws or a direct action on one or more mediator receptors. The formalin model alone cannot be used to ascertain the anti-inflammatory effects of the Citrus species. We therefore used the carrageenan-induced inflammation in a subcutaneous air pouch SAP model. This model is characterized by a drastic increase in leukocytes, cytokines, inflammatory mediators and protein after the carrageenan injection [ 20 ]. The mechanism by which carrageenan induces the inflammatory response is complex and involves the liberation of several mediators and an increase in vascular permeability. Our results indicate that EOs induce an anti-inflammatory effect by reducing several of the parameters observed in carrageenan-induced inflammation. These results cannot guarantee the exact local action of the EOs, but do suggest that EOs can be acting in one or more mediator systems involved in the inflammation. During the inflammatory event there is also an increase in cytokine production. Tumor necrosis factor-α TNF-α , interleukin-1β IL-1β , and interferon-γ IFN-γ have important roles in the maintenance of the inflammatory profile [ 26 , 27 ]. Our results indicate that EOs have significant anti-inflammatory effects in reducing all parameters evaluated. Since IL-1β and TNF-α regulate leukocyte migration, we can infer that a reduction in leukocyte number may have been a direct effect of EOs as well as an indirect effect resulting from the reduction in the levels of those cytokines. Taken together, these results indicate that these EOs act similarly to immunomodulators in reducing cell migration and inflammatory mediator production. Our results are in agreement with others that showed various essential oils or their constituents inhibit cytokine production. For example, 1,8-cineol inhibited TNF-α and IL-1β in human lymphocytes, α-humulene reduced TNF-α production, and terpinenol suppressed the production of TNF-α, IL-1β, IL-8, IL, and PGE2 by LPS-activated monocytes [ 28 ]. latifolia reduced TNF-α and IL levels in zymosan-induced peritonitis [ 9 ]. Despite all EOs present limonene in there constitution a difference occurred between the inhibitions observed in the formalin-induced licking and carrageenan-induced cell migration models. A possible explanation to the differences could be the fact that both models are very different. In the first one we observe liberation of inflammatory mediators such as histamine, serotonin and bradykinin in mice paws [ 17 ]. In the second model we have the involvement of several other mediators and systems, liberation of nitrogen reactive species ROS and cytokines [ 19 ]. Additionally, C. aurantifolia EO demonstrated some toxicity, likely due to the presence of high amounts of citral. Others have demonstrated a toxic effect of citral. However, those groups used different models such as embryofetal [ 29 ] and carcinogenesis induction [ 30 ]. |
| Thieme E-Books & E-Journals - | Nogueira AC, Carvalho RR, Souza CA, Chahoud I, Paumgartten FJ. Int Rev Neurobiol. Lota ML, Serra DR, Tomi FL, Jacquemond C, Casanova J. New research suggests that running may not aid much with weight loss, but it can help you keep from gaining weight as you age. Sufficient data indicate the prevention and treatment activities of CACH on liver diseases are mainly achieved through anti-inflammatory, antioxidant and intestinal microflora regulation. Synthesis of limonoid natural products eur. |
| Anti-inflammatory Effect of Essential Oil from Citrus aurantium L. var. amara Engl | Compound identification The identification of individual components was based on i a comparison of the mass spectral fragmentation patterns with those stored in the NIST Mass Spectral Library and ii a comparison of the GC Kovats retention indices KI on a DB-5 column with those of authentic compounds and from literature data [ 13 ]. A new HPLC method developed for the analysis of oxygen heterocyclic compounds in Citrus essential oils. It was also observed that C. Publication types Research Support, Non-U. There are two phases, with the first neurogenic phase occurring peripherally and resulting from formalin activation of nociceptors located in the tissue and the second inflammatory phase occurring after the release of a multitude of molecules, such as histamine, serotonin, and bradykinin, developing an inflammatory response [ 25 ]. For example, it was found that CACH contains 16 kinds of amino acids, including eight kinds of essential amino acids, and the content of ASP is the highest Zheng et al. |
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