Regulation of Nrf2 and NF-κB activities may contribute to the anti-inflammatory mechanism of xylopic acid.

Xylopic acid (XA) is a kaurene diterpene which naturally exists in African plants such as Xylopia aethiopica. It has been established to exhibit acute and chronic anti-inflammatory activities from our earlier studies. This current work sets out to shed light on the potential molecular target(s) of xylopic acid. Selection of investigated targets (NF-κB, Nrf2 and PTP1B) was based on an unbiased approach, using the SPiDER in silico prediction tool, and a candidate approach, examining well-known anti-inflammatory targets. Reporter gene assays were used to test for altered NF-κB and Nrf2 activities in transfected HEK or CHO cells, respectively, and immunoblot and flow cytometric analyses examined protein expression of the Nrf2/NF-kB target genes HO-1 and VCAM-1 in HUVEC. An effect of XA on PTP1B activity assay was studied using an in vitro enzyme assay with recombinant human enzyme and pNPP as substrate as well as by looking at insulin receptor phosphorylation in HepG2 cells. XA at 30 µM significantly (p < 0.001) inhibited the NF-κB-dependent reporter gene expression and enhanced activation of Nrf2 in a concentration-dependent manner when compared to the control. XA also marginally increased HO-1 protein expression levels while expression of VCAM-1 was reduced to 70% in XA-treated endothelial cells. However, XA did not show any sign of inhibition of PTP1B or a related phosphatase. Our findings suggest that the anti-inflammatory mechanism of XA entails the inhibitory effect on NF-κB and an increased activity of Nrf2, accompanied by increased expression of HO-1 and reduced expression of VCAM-1.

Hydro-ethanolic leaf extract of Ziziphus abyssinica Hochst Ex A. Rich (Rhamnaceae) exhibits anti-nociceptive effects in murine models.

BACKGROUND: Despite substantial advances in pain research and treatment, millions of people continue to suffer from pain and this has been attributed mainly to the unavailability of effective and safer analgesics. The use of plants as medicines is still widespread and plants constitute a large source of novel phytocompounds that might become leads for the discovery of newer, effective and safer alternatives. Various parts of Ziziphus abyssinica have been used in folk medicine in several African countries as painkillers. However, there is no report on the possible anti-nociceptive effects of this plant especially the leaves, hence the need for this current study. METHODS: The possible anti-nociceptive activity of hydro-ethanolic leaf extract of Ziziphus abyssinica (EthE) was assessed in rodents using chemical (acetic acid, formalin and glutamate), thermal (tail-immersion test) and mechanical/inflammatory (carrageenan) models of nociception. RESULTS: EthE (30-300 mg/kg, p.o.) dose-dependently and significantly inhibited chemical-induced nociception with a maximum inhibition of 86.29 ± 2.27%, 76.34 ± 5.67%, 84.97 ± 5.35%, and 82.81 ± 5.97% respectively for acetic acid, formalin (phase 1), formalin (phase 2) and glutamate tests at its highest dose. EthE also dose-dependently and significantly increased reaction times in both tail-immersion and carrageenan-induced hypernociceptive tests. The activities of the extract in the various models were comparable with the effect of morphine hydrochloride and diclofenac sodium used as standard analgesic drugs. CONCLUSION: Oral administration of hydro-ethanolic leaf extract of Ziziphus abyssinica ameliorates nocifensive behaviours associated with chemical-, thermal- and mechanical/inflammatory - induced nociceptive pain.

Possible mechanisms involved in the anti-nociceptive effects of hydro-ethanolic leaf extract of Ziziphus abyssinica.

CONTEXT: Various parts of Ziziphus abyssinica Hochst ex. A. Rich (Rhamnaceae) have been used in Ghanaian and African traditional medicine as an analgesic. However, there are little scientific data to support the anti-nociceptive effects of the hydro-ethanolic leaf extract of Ziziphus abyssinica (EthE) as well as the possible mechanisms involved in its anti-nociceptive effects. PURPOSE: To predict possible nociceptive pathways involved in the anti-nociceptive effects of EthE. MATERIALS AND METHODS: The effect of EthE (30, 100 and 300 mg/kg) on intraplantar injection of pain mediators such as interleukin-1ß, tumour necrosis factor-α, prostaglandin E2 and bradykinin was evaluated in male Sprague Dawley rats using Randall-Selitto test for 5 h. The effect of specific antagonists to the opioidergic, adenosinergic, ATP-sensitive K+ channels, nitric oxide, serotonergic, muscarinic, adrenergic and voltage-gated calcium channel on the anti-nociceptive effect of EthE (100 mg/kg) was evaluated using the formalin test in male imprinting control region (ICR) mice for 1 h. RESULTS: Pretreatment of the rats with EthE significantly reversed the hypernociception induced by intraplantar injection of TNF-α (F4,120 = 10.86, p < 0.0001), IL-1ß (F4,120 = 14.71, p < 0.0001), bradykinin (F4,80 = 12.52, p < 0.0001) and prostaglandin E2 (F5,144 = 6.165, p = 0.0001). The anti-nociceptive effect exhibited by EthE in the formalin test was reversed by systemic administration of NG-l-nitro-arginine methyl ester, naloxone, theophylline and glibenclamide. CONCLUSIONS: EthE inhibits hypernociception induced by TNF-α, IL-1ß, bradykinin and prostaglandin E2. EthE exhibited anti-nociceptive effects possibly mediated through opioidergic, adenosinergic, ATP-sensitive potassium channels and nitric oxide cyclic GMP pathways.

An evaluation of the anti-inflammatory, antipyretic and analgesic effects of hydroethanol leaf extract of Albizia zygia in animal models.

CONTEXT: The leaves of Albizia zygia (DC.) J.F. Macbr. (Leguminosae-Mimosoideae) are used in Ghanaian traditional medicine for the treatment of pain, inflammatory disorders and fever (including malaria). OBJECTIVES: The present study evaluated the anti-inflammatory, antipyretic and analgesic effects of the hydroethanol leaf extract of Albizia zygia (AZE) in animal models. MATERIALS AND METHODS: The anti-inflammatory and antipyretic effects of AZE were examined in the carrageenan-induced foot oedema model and the baker's yeast-induced pyrexia test respectively. The analgesic effect and possible mechanisms of action were also assessed in the formalin test. RESULTS: AZE (30-300 mg/kg, p.o.), either preemptively or curatively, significantly inhibited carrageenan-induced foot edema in 7-day-old chicks (ED50 values; preemptive: 232.9 ± 53.33 mg/kg; curative: 539.2 ± 138.28 mg/kg). Similarly, the NSAID diclofenac (10-100 mg/kg, i.p.) significantly reduced the oedema in both preemptive (ED50: 21.16 ± 4.07 mg/kg) and curative (ED50: 44.28 ± 5.75 mg/kg) treatments. The extract (30-300 mg/kg, p.o.) as well as paracetamol (150 mg/kg, p.o.) also showed significant antipyretic activity in the baker's yeast-induced pyrexia test (ED50 of AZE: 282.5 ± 96.55 mg/kg). AZE and morphine (1-10 mg/kg, i.p.; positive control), exhibited significant analgesic activity in the formalin test. The analgesic effect was partly or wholly reversed by the systemic administration of naloxone, theophylline and atropine. CONCLUSION: The results suggest that AZE possesses anti-inflammatory, antipyretic and analgesic properties, which justifies its traditional use. Also, the results show the involvement of the opioidergic, adenosinergic and the muscarinic cholinergic pathways in the analgesic effects of AZE.

Effects of an ethanol extract and the diterpene, xylopic acid, of Xylopia aethiopica fruits in murine models of musculoskeletal pain.

CONTEXT: Fruits of Xylopia aethiopica (Dunal) A. Rich. (Annonaceae) are used traditionally to manage arthritis, headache and other pain disorders. OBJECTIVE: The analgesic properties of the X. aethiopica ethanol fruit extract (XAE) and xylopic acid (XA) were evaluated in musculoskeletal pain models. MATERIALS AND METHODS: Acute muscle pain was induced in gastrocnemius muscle of Sprague-Dawley rats with 3% carrageenan (i.m.). Rats received XAE (30-300 mg/kg), XA (10-100 mg/kg) or morphine (1-10 mg/kg) after 12 h. Effects of XAE and XA on muscle pain were assessed by measuring post-treatment grip strength of the rats. Chronic muscle pain was similarly induced, but drug treatment was on the eighth day and effects of XAE and XA assessed with Randall-Selitto test for hyperlagesia. Acute-skeletal pain was induced in knee joints of rats with 3% carrageenan-kaolin mixture and effects determined 12-h later. Similar induction protocol was used for chronic knee pain with treatment and measurement as done for chronic muscle pain. RESULTS: XAE and XA significantly and dose-dependently ameliorated both acute muscle (ED50 mg/kg: XAE = 22.9; XA = 6.2) and skeletal hyperalgesia (XAE = 39.9; XA = 17.7) induced by 3% carrageenan. Similarly, chronic skeletal hyperalgesia was reduced by XAE and XA treatment similar to morphine (ED50: XAE = 13.0; XA = 4.6). This reduction was also seen in chronic muscle hyperalgesia (ED50: XAE = 79.1; XA = 42.7). XAE and XA significantly reduced the spread of hyperalgesia to contralateral limbs in both models of chronic hyperalgesia. CONCLUSION: These findings establish analgesic properties of the ethanol fruit extract of X. aethiopica and xylopic acid in musculoskeletal pain.

Synergistic antidepressant-like effect of xylopic acid co-administered with selected antidepressants.

Background: Xylopic acid (XA), a kaurene diterpene from the dried fruits of Xylopia aethiopica, has anxiolytic- and antidepressant-like activity in mice and zebrafish. We aimed to assess the potential synergistic antidepressant-like effects of XA when combined with selected antidepressants in the mouse forced-swim test. Materials and methods: The antidepressant-like effect of xylopic acid (XA) (10, 30, 100 mgkg-1), fluoxetine (Flx) (3, 10, 30 mgkg-1), sertraline (Sert) (3, 10, 30 mgkg-1), imipramine (Imi) (10, 30, 100 mgkg-1) and ketamine (Ket) (0.1, 0.3, 1.0 mgkg-1), was evaluated in forced swim test. The dose (ED50) that achieved a 50% reduction in immobility time was determined from the respective log-dose response curves. XA and the selected antidepressants were co-administered in fixed-dose ratio combinations (1/2:1/2, 1/4:1/4, 1/8:1/8) of the ED50 to identify the experimental ED50 (ED50mix). The theoretical ED50(ED50add), of all combinations was determined using isobolograms and compared with the ED50mix to identify the nature of the interaction. The effect of dose combinations on general locomotor activity was assessed in the open-field test. Results: The interaction index (γ) for the following XA combinations, XA/Flx, XA/Sert, XA/Imi and XA/Ket were 0.42, 0.41, 0.31 and 0.34. An independent sample t-test revealed that the experimental ED50 (ED50mix) was significantly lower than the theoretical ED50 (ED50add) in all combinations of XA, indicative of a synergistic antidepressant-like effect. However, combinations of XA with ketamine significantly reduced general locomotor activity at all dose combinations. Conclusion: The co-administration of xylopic acid and fluoxetine, imipramine, sertraline and ketamine produces a synergistic antidepressant-like effect in mice.

Anti-Arthritic Effect of the Hydroethanolic Root Extract of Psydrax subcordata in Rats.

Background: In Ghana, decoctions of various parts of Psydrax subcordata, Bridson (Rubiaceae) are employed in the management of inflammatory conditions. However, not much scientific data is available to back such folkloric use of the plant. This study, therefore, seeks to investigate the chronic anti-inflammatory activity of hydroethanolic root extract of Psydrax subcordata (PSRE) using the adjuvant-induced arthritis model in rats. Methods: Freund's adjuvant-induced arthritis model was used to assess the ameliorative effects of PSRE in chronic inflammation. The effect of PSRE on tissue and joint integrity in arthritis was also evaluated by histopathology and microscopy. The effect of PSRE on oxidative markers and serum transforming growth factor (TGF) beta 1 was also determined via chemical assays. Results: Oral PSRE (30-300 mg/kg) inhibited both ipsilateral and contralateral paw arthritis when given prophylactically and therapeutically in rats. It reduced paw defect on X-ray with histologically-reduced inflammatory cells and synovial hyperplasia. Finally, PSRE significantly reduced TGF-beta 1 levels and raised antioxidants such as reduced glutathione, catalase, and superoxide dismutase levels in arthritic rats. Conclusion: The findings show that hydroethanolic root extract of Psydrax subcordata possesses anti-inflammatory properties in rodents.

Antidepressant-like effect of Albizia zygia root extract in murine models.

Background Albizia zygia (DC.) J.F. Macbr. (Leguminosae) has been used to treat mental disorders in traditional African medicine. Nonetheless, there is limited scientific evidence to justify its present use. The aim of this study was to evaluate the antidepressant activity of the hydroethanolic extract of A. zygia roots (AZE) in murine models. Methods AZE was evaluated in the tail suspension test, forced swim test, and the repeated open-space swim test of depression. In order to elucidate the mechanisms of action, the activity of AZE was re-evaluated after treating mice with selective inhibitors of monoamine biosynthesis. The potential of AZE to influence spontaneous locomotion was also examined. Results AZE (100-1000 mg/kg, p.o.) reduced the immobility time of mice in the tail suspension and forced swim tests (at least p < 0.05). In the repeated open-space swim test, AZE reduced the immobility time (at least p < 0.05) while concomitantly increasing the distance swam by mice (p < 0.01). However, the antidepressant-like activity of AZE was attenuated by α-methyl-para-tyrosine and reserpine (p < 0.0001) but not para-chlorophenylalanine. Conclusions The results of this study indicate that AZE possesses antidepressant-like properties and support the traditional use of AZE for the treatment of depression.

Albizia zygia root extract exhibits antipsychotic-like properties in murine models of schizophrenia.

BACKGROUND: The root extract of Albizia zygia (DC.) J.F. Macbr. (Leguminosae) is used to manage mental disorders in African traditional medicine. However, its value, particularly, against negative and cognitive symptoms of schizophrenia have not been evaluated. AIM: The aim of this study was to evaluate the antipsychotic properties of the hydroethanolic root extract of Albizia zygia (AZE) against positive, negative and cognitive symptoms of schizophrenia in animal models. MATERIALS AND METHODS: The effects of AZE (30-300 mg kg-1) were evaluated against apomorphine-induced cage climbing as well as ketamine -induced hyperlocomotion, -enhanced immobility, -impaired social interaction and novel object recognition. The propensity of AZE to induce catalepsy and to attenuate haloperidol-induced catalepsy were also investigated. RESULTS: AZE 30-300 mg kg-1 significantly reduced apomorphine-induced climbing behaviour as well as ketamine-induced hyperlocomotion, immobility and object recognition deficits (at least P < 0.05). Moreover, the extract showed no cataleptic effect but significantly inhibited haloperidol-induced catalepsy at a dose of 30 mg kg-1 (P < 0.05). CONCLUSION: The root extract of Albizia zygia exhibited an antipsychotic-like activity in mice with potential to alleviate positive, negative and cognitive symptoms of schizophrenia.

Anticonvulsant effect of the hydroethanolic leaf extract of Psydrax subcordata (DC.) Bridson in murine models.

ETHNOPHARMACOLOGICAL RELEVANCE: Psydrax subcordata (DC.) Bridson is a tropical medicinal plant used traditionally for the management of epilepsy. However, there is little scientific evidence to support its use. AIM OF STUDY: The current study investigated the anticonvulsant properties of the hydroethanolic leaf extract of Psydrax subcordata (PSE) in animal models. MATERIALS AND METHODS: The anticonvulsant effects were evaluated in mouse models of acute seizures (pentylenetetrazole-, picrotoxin-, 4-aminopyridine-, strychnine- and maximal electroshock-induced seizure tests) and status epilepticus (Lithium/pilocarpine-induced SE). The role of GABAergic mechanisms in the actions of the extract was also examined by pre-treatment of animals with flumazenil in the pentylenetetrazole test. RESULTS: The extract (30, 100 and 300mg/kg, p.o.) significantly delayed the onset and decreased the duration and frequency of pentylenetetrazole- and picrotoxin-convulsions. PSE also reduced the duration of tonic hind limb extensions in the maximal electroshock-induced seizure test. Furthermore, PSE pre-treatment significantly delayed the onset of seizures and improved survival in the 4-aminopyridine-induced seizure test. In the strychnine-induced seizure test, PSE treatment did not significantly affect the latency to convulsions and time until death when compared to controls. PSE exhibited anticonvulsant effects in the lithium/pilocarpine test by delaying the onset of seizures and status epilepticus as well as reducing the severity of seizures and mortality of mice. Again, the anticonvulsant effect of PSE (100mg/kg, p.o.) was blocked by pre-treatment with flumazenil in the PTZ test. CONCLUSION: PSE has anticonvulsant activity in animal models, and this effect may be mediated, at least partly, through GABAergic mechanisms.

An isobolographic analysis of the anti-nociceptive effect of geraniin in combination with morphine or diclofenac.

BACKGROUND: Geraniin, a dehydroellagitannin, is a major component of the aqueous extract of the aerial parts of Phyllanthus muellerianus (Kuntze) Exell. (Euphorbiaceae). Several Phyllanthus species are traditionally used for painful disorders. The anti-nociceptive effects of the aqueous extract of the aerial parts of P. muellerianus and of geraniin have been scientifically established. The aim of the paper is to determine whether a combination of geraniin and diclofenac or geraniin and morphine leads to better anti-nociceptive effects. METHODS: The nature of the interactions of morphine and diclofenac with geraniin was evaluated by undertaking the isobolographic analysis. Mice were treated with geraniin (3-30 mg/kg), morphine (1-10 mg/kg), and diclofenac (10-100 mg/kg) to obtain the ED50 values of the agents in the formalin test. Dose-response curves were then obtained and analyzed after the co-administration of geraniin with morphine or diclofenac in fixed ratio (1:1) combinations based on specific fractions (1/2, 1/4, and 1/8) of their respective ED50 values for the formalin test. RESULTS: Geraniin was less potent than morphine but more potent than diclofenac in the formalin-induced nociception. The isobolographic analysis of geraniin/morphine (G/M) and geraniin/diclofenac combinations (G/D) at different fractions revealed the potentiation of their anti-nociceptive effects. The degrees of potentiation, which were calculated as interaction indices, showed synergism for both combinations in both phase I (G/M: 0.040, G/D: 0.017) and phase II (G/M: 0.004, G/D: 0.002) of the formalin test. CONCLUSIONS: The present study demonstrates synergism for the co-administration of geraniin with both morphine and diclofenac.

Albizia zygia (DC.) J.F. Macbr. (Leguminosae-Mimosoideae) root extract exhibits anti-nociceptive and antipyretic activities in murine models.

ETHNOPHARMACOLOGICAL RELEVANCE: The root extract of Albizia zygia (DC.) J.F. Macbr. (Leguminosae-Mimosoideae) is traditionally used in the management of pain and fever. However, little scientific data exists in literature to support its use. AIM OF STUDY: The present study evaluated the anti-nociceptive and antipyretic properties of the hydroethanolic extract of the roots of Albizia zygia in animal models. MATERIALS AND METHODS: The analgesic effects were investigated in chemical (acetic acid-induced abdominal writhing and formalin tests), thermal (tail-immersion test) and mechanical (carrageenan-induced hyperalgesia) pain models. Possible mechanisms of anti-nociception were also assessed with antagonists in the formalin test. The anti-pyretic effect was evaluated using the baker yeast-induced pyrexia model in young rats. RESULTS: The extract (30-300mg/kg, p.o.) and positive controls, diclofenac (3-30mg/kg, i.p.) and morphine (1-10mg/kg, i.p.), significantly (at least P<0.01) attenuated acetic acid-induced visceral pain, formalin- induced paw pain (both neurogenic and inflammatory), thermal pain as well as carrageenan-induced mechanical hyperalgesia in animals. The anti-nociceptive effect of the extract was reversed (at least P<0.05) by the pre-emptive administration of naloxone and atropine; the administration of theophylline, however, exhibited no significant (P>0.05) inhibition of anti-nociception. The extract (30-300mg/kg, p.o) and paracetamol (15-150mg/kg, p.o.) both reversed yeast-induced pyrexia in rats with ED50 values of 48.59±2.59 and 26.19±1.33mg/kg respectively. CONCLUSION: The findings indicate that the extract possesses significant anti-nociceptive and antipyretic effects which justify its traditional use in the management of pain and fever. Also, anti-nociceptive effect of the extract involves opioidergic and muscarinic cholinergic mechanisms.

Anti-inflammatory activity of aqueous leaf extract of Phyllanthus muellerianus (Kuntze) Exell. and its major constituent, geraniin.

ETHNOPHARMACOLOGICAL RELEVANCE: Phyllanthus muellerianus (Kuntze) Exell. which belongs to the Family Euphorbiaceae is a shrub widely distributed in West Africa. It is used traditionally to manage wounds and wound infections, menstrual disorders, fevers, pain and inflammation. Hence to confirm its ethnobotanical uses in managing inflammation, we investigated the anti-inflammatory properties of aqueous leaf extract of P. muellerianus (PLE) and its major isolate, geraniin in experimentally-induced inflammation in rats. MATERIALS AND METHODS: Carrageenan induced oedema and adjuvant induced arthritis models in rats were used in this study. RESULTS: In the carrageenan-induced acute inflammation, both 300mg/kg PLE-treated and 10mg/kg geraniin-treated groups significantly reduced the mean maximal swelling attained at 4h to 46.75±4.97% (p<0.01) and 61.65±6.70% (p<0.05), respectively, from the inflamed control response of 122.60±16.39%. In the adjuvant-induced chronic inflammation, both PLE-treated (100 and 300mg/kg) groups and geraniin-treated (10 and 30mg/kg) groups significantly (p<0.001) reduced the total limb swelling over 16 days in the polyarthritic phase compared to the arthritic control. These observations were supported by the radiograph records and the histological investigations of the hind limbs which showed reduced bone damage in both PLE and geraniin-treated rats. CONCLUSION: The findings may confirm the ethnobotanical use of PLE in the management of inflammatory disorders or conditions and observed anti-inflammatory property of PLE may largely be due to its major constituent, geraniin.

An isobolographic analysis of the antinociceptive effect of xylopic acid in combination with morphine or diclofenac.

BACKGROUND: A common practice of managing pain globally is the combination of analgesics and this is aimed at facilitating patient compliance, simplifying prescription, and improving efficacy without increasing adverse effects. Fruit extracts of Xylopia aethiopica are used traditionally in the management of pain disorders and xylopic acid (XA) present in the fruit extract have been shown to possess analgesic properties in animals. There is the likelihood of concomitant use of XA and the commonly used analgesics in traditional settings. This study, therefore, evaluated the pharmacologic interaction between XA/morphine and xylopic/diclofenac combinations. METHODS: The formalin test and acetic acid writhing test were used to study the antinociceptive activity of XA, morphine, and diclofenac. The isobolographic analysis was used to study the antinociceptive interactions between XA co-administered with morphine or diclofenac. RESULTS: Results obtained revealed that XA (10-100 mg/kg), morphine (1-10 mg/kg), and diclofenac (1-10 mg/kg) produced dose-related antinociception with different potencies in the formalin and acetic acid writhing tests. Isobolographic analysis of XA/morphine and XA/diclofenac combinations revealed potentiation of their antinociceptive effects. The degree of potentiation calculated as interaction index showed synergism for both combinations in all the nociceptive tests. CONCLUSION: In conclusion, the present study demonstrated synergism for the co-administration of XA with morphine or diclofenac.

Analgesic effects of stem bark extracts of Trichilia monadelpha (Thonn.) JJ De Wilde.

OBJECTIVES: Various parts of Trichilia monadelpha (Thonn) JJ De Wilde (Fam. Meliaceae) are used in Ghanaian traditional medicine for the treatment of painful and inflammatory conditions. The present study examined the analgesic properties of the petroleum ether (PEE), ethyl acetate (EAE), and the hydro-ethanolic (HAE) extract of the stem bark of the plant in murine models. MATERIALS AND METHODS: PEE, EAE, and HAE were assessed in chemical (acetic acid-induced abdominal writhing and formalin tests), thermal (hot plate test), and mechanical (Randall-Selitto paw pressure test) pain models. The possible mechanisms of the antinociceptive action were also examined with various antagonists in the formalin test. RESULTS: HAE, EAE, and PEE, each at doses of 10-100 mg/kg orally, and the positive controls (morphine and diclofenac) elicited significant dose-dependent antinociceptive activity in the chemical (acetic acid abdominal writhing and formalin tests), thermal (hot plate test), and mechanical (Randall-Selitto paw pressure test) pain models in rodents. The antinociceptive effect of HAE was partly or wholly reversed by systemic administration of atropine, naloxone, and glibenclamide. The antinociceptive effects of EAE and PEE were inhibited by atropine. CONCLUSION: The extracts HAE, EAE, and PEE caused dose-related antinociception in chemical, thermal, and mechanical models of pain in animals. The mechanism of action of HAE involves an interaction with muscarinic cholinergic, adenosinergic, opioidergic pathways, and ATP-sensitive K+ channels while that of EAE and PEE involve the muscarinic cholinergic system.