Bronchodilator Activity of Aerial Parts of Polygonatum verticillatum Augmented by Anti‐inflammatory Activity: Attenuation of Ca2+ Channels and Lipoxygenase

Polygonatum verticillatum is commonly used for the treatment of asthma and inflammation. The current study was aimed to scrutinize the pharmacological profile of methanolic extract of the aerial parts (PA). Isolated tracheal preparations were used for the evaluation of bronchodilatory activity, whilst the in vivo carrageenan‐induced paw oedema test and an in vitro lipoxygenase (LOX) inhibitory assay were used for the assessment of the anti‐inflammatory profile of PA. When tested against carbachol and K⁺ (80 mM)‐induced contractions, PA caused complete inhibition of isolated rabbit tracheal preparations in a dose‐dependent mode, similar to verapamil. While elucidating possible mechanism, PA shifted the Ca²⁺ concentration–response curves to the right, analogous to that produced by verapamil, confirming a Ca²⁺ channel blocker‐like activity. PA provoked profound reduction in paw oedema with a maximum protection of 60.87% at 200 mg/kg i.p. in a dose‐dependent manner which was augmented by its prominent LOX inhibitory activity (IC₅₀: 125 µg/mL). These findings authenticated its therapeutic potential in the treatment of asthmatic and inflammatory conditions. Copyright © 2012 John Wiley & Sons, Ltd.     

Anti‐Inflammatory Sesquiterpene Lactones from Lychnophora trichocarpha Spreng. (Brazilian Arnica)

The aerial parts of Lychnophora trichocarpha Spreng. (Asteraceae) are used macerated in water or ethanol to treat inflammation, pain, rheumatism, contusions, bruises and insect bites in Brazilian traditional medicine. In this study, anti‐inflammatory activity of ethanol extract from aerial parts of L. trichocarpha and its ethyl acetate fraction was investigated. Sesquiterpene lactones, lychnopholide (Lyc) and eremantholide C (EreC), isolated of ethyl acetate fraction, were also assayed for in vitro and in vivo anti‐inflammatory activity. Topical treatment with ointments containing ethanol extract, its ethyl acetate fraction and sesquiterpene lactones significantly reduced carrageenan‐induced mice paw oedema. In vitro assays demonstrated that Lyc inhibited interferon ‐γ/lipopolysaccharide ‐stimulated nitric oxide (NO) production in J774A.1 macrophages and increased production of IL‐10 anti‐inflammatory cytokine. The reduction of tumor necrosis factor‐α (TNF‐α) production by EreC was accompanied by an increased production of IL‐10 in a concentration‐dependent manner in J774A.1 macrophages. The anti‐inflammatory effect of Lyc seems to involve the inhibition of production of NO and increased production of IL‐10. The mechanism of the effect of EreC on the reduction of carrageenan‐induced paw oedema may be attributed to inhibition of production of TNF‐α and stimulation of IL‐10 production. The results corroborate the use of ethanol extract from Lychnophora trichocarpha in folk medicine for anti‐inflammatory action and indicate that the topical route is suitable for use. Copyright © 2012 John Wiley & Sons, Ltd.     

Houttuynia cordata Thunb. Volatile Oil Exhibited Anti‐inflammatory Effects In Vivo and Inhibited Nitric Oxide and Tumor Necrosis Factor‐α Production in LPS‐stimulated Mouse Peritoneal Macrophages In Vitro

Houttuynia cordata Thunb. (HC) is a medicinal herb that generally used in traditional Chinese medicine for treating allergic inflammation. The present study investigated the inhibitory effect of the volatile oil from HC Thunb. on animal models of inflammation and the production of inflammatory mediators in vivo and in vitro. In vivo, xylene‐induced mouse ear edema, formaldehyde‐induced paw edema and carrageenan‐induced mice paw edema were significantly decreased by HC volatile oil. HC volatile oil showed pronounced inhibition of prostaglandin (PG) E₂ and malondialdehyde production in the edematous exudates. In vitro exposure of mouse resident peritoneal macrophages to 1, 10, 100 and 1000 µg/mL of HC volatile oil significantly suppressed lipopolysaccharide (LPS)‐stimulated production of NO and tumor necrosis factor‐α (TNF‐α) in a dose‐dependent manner. Exposure to HC volatile oil had no effect on cell viability and systemic toxicity. Furthermore, HC volatile oil inhibited the production of NO and TNF‐α by down‐regulating LPS‐stimulated iNOS and TNF‐α mRNA expression. Western blot analysis showed that HC volatile oil attenuated LPS‐stimulated synthesis of iNOS and TNF‐α protein in the macrophages, in parallel. These findings add a novel aspect to the biological profile of HC and clarify its anti‐inflammatory mechanism. Copyright © 2012 John Wiley & Sons, Ltd.     

The phytochemical and anti‐inflammatory studies of Dillenia suffruticosa leaves

The Dillenia suffruticosa leaves (Dilleniaceae), a folk medicine recommended in Southeast Asia for treating inflammation, were phytochemically studied for the first time and assessed for suppression of λ‐carrageenan‐induced paw oedema in rats. The crude methanolic extract orally administered at 5,000 mg/kg, displayed no toxicity and at 250 to 1,000 mg/kg significantly suppressed the paw oedema. Two‐isolated triterpenoids, betulinic acid (1) and koetjapic acid (2) orally administered at 50 mg/kg, significantly reduced the paw oedema, (p < 0.001) and (p < 0.005) at the fourth h onwards to 47.36% ± 2.23 and 53.43% ± 7.09, respectively, from 95.90% ± 6.88 oedema induced by λ‐carrageenan alone. 1 and the isolated flavonoids of vitexin (3), tiliroside (4), and kaempferol (5), displayed moderately more of cyclooxygenase (COX)‐2 than COX‐1 enzyme inhibition, whereas 2 was slightly more inhibition of COX‐1. The in silico molecular docking studies provided support to the in vitro COX studies that the isolated compounds formed H‐bonding with the amino acid residues at the COX‐2 catalytic sites. The triterpenoids were bound to the peroxidase, possibly inhibiting the peroxidase reaction, whereas the flavonoids interacted more at the cyclooxygenase, resembling celecoxib, therefore providing evidences that these compounds were responsible for the anti‐inflammatory properties of D. suffruticosa.     

Anti‐inflammatory properties of cryptolepine

Cryptolepine is the major alkaloid of the West African shrub, Cryptolepis sanguinolenta. Cryptolepine has been shown to inhibit nitric oxide production, and DNA binding of Nuclear Factor‐kappa B following inflammatory stimuli in vitro. In order to validate the anti‐inflammatory property of this compound in vivo, we investigated its effects on a number of animal models of inflammation. Cryptolepine (10–40 mg/kg i.p.) produced significant dose‐dependent inhibition of the carrageenan‐induced rat paw oedema, and carrageenan‐induced pleurisy in rats. These effects were compared with those of the non‐steroidal anti‐inflammatory drug indomethacin (10 mg/kg). At doses of 10–40 mg/kg i.p., cryptolepine inhibited lipopolysaccharide (LPS)‐induced microvascular permeability in mice in a dose‐related fashion. Oral administration of up to 40 mg/kg of the compound for four consecutive days did not induce gastric lesion formation in rats. Analgesic activity was also exhibited by cryptolepine through a dose‐related (10–40 mg/kg i.p.) inhibition of writhing induced by i.p. administration of acetic acid in mice. The results of this study reveal that cryptolepine possesses in vivo anti‐inflammatory activity. Copyright © 2009 John Wiley & Sons, Ltd.     

Anti‐Inflammatory and Antinociceptive Activity of Urera aurantiaca

Urera aurantiaca Wedd. (Urticaceae) is a medicinal plant commonly used in traditional medicine to relieve pain in inflammatory processes. In the present study, the in vivo anti‐inflammatory and antinociceptive effects of U. aurantiaca methanolic extract and its possible mechanisms of action were investigated. The extract showed anti‐inflammatory activity in the ear edema in mice test (34.3% inhibition), myeloperoxidase (MPO) activity was markedly reduced in animals administered with the extract: within 49.6% and 68.5%. In the histological analysis, intense dermal edema and intense cellular infiltration of inflammatory cells were markedly reduced in the ear tissue of the animals treated with the extract. In the carrageenan‐induced hind paw edema in rats assay the extract provoked a significant inhibition of the inflammation (45.5%, 5 h after the treatment) and the MPO activity was markedly reduced (maximum inhibition 71.7%), The extract also exhibited significant and dose‐dependent inhibitory effect on the increased vascular permeability induced by acetic acid. The extract presented antioxidant activity in both 2,2‐diphenyl‐1‐picrylhydrazyl and 2,2′‐azinobis 3‐ethylbenzothiazoline 6‐sulfonic acid tests and its total phenol content was 35.4 ± 0.06 mg GAE/g of extract. Also, the extract produced significant inhibition on nociception induced by acetic acid (ED₅₀: 8.7 mg/kg, i.p.) administered intraperitoneally and orally. Naloxone significantly prevented this activity. Copyright © 2014 John Wiley & Sons, Ltd.     

Anti‐allergic activity of principles from the roots and heartwood of caesalpinia sappan on antigen‐induced β‐hexosaminidase release

The dichloromethane extract of the roots and heartwood of Caesalpinia sappan exhibited potent inhibitory activity against β‐hexosaminidase release as marker of degranulation in rat basophilic leukemic (RBL‐2H3) cells, with inhibition of 98.7% and 87.5% at concentration of 100 µg/ml, respectively. These extracts were further separated by chromatographic techniques to give two chalcones and seven homoisoflavones. Among the compounds tested, sappanchalcone (2) possessed the most potent effect against allergic reaction in RBL‐2H3 cells with an inhibitory concentration (IC₅₀) value of 7.6 µM, followed by 3‐deoxysappanchalcone (1, IC₅₀ = 15.3 µM), whereas other compounds showed moderate and mild effects. The results suggested the following structural requirements of chalcones (1 and 2) and homoisoflavones (3‐9) for anti‐allergic activity: (i) chalcone exhibited higher activity than homoisoflavone (ii) vicinal hydroxylation at B‐ring of chalcone conferred higher activity than one hydroxylation; and (iii) for homoisoflavone, the hydroxyl groups at C‐3 and C‐4 positions decreased the activity. This is the first report of C. sappan for anti‐allergic activity. Copyright © 2009 John Wiley & Sons, Ltd.     

Picrorhiza kurroa Inhibits Experimental Arthritis Through Inhibition of Pro‐inflammatory Cytokines,Angiogenesis and MMPs

The present study investigates the anti‐arthritic activity of Picrorhiza kurroa (PK), on formaldehyde and adjuvant‐induced arthritis (AIA) in rat. Administration of Picrorhiza kurroa rhizome extract (PKRE) significantly inhibited joint inflammation in both animal models. In AIA‐induced arthritic rat, treatment with PKRE considerably decreased synovial expression of interleukin‐1β (IL‐1β), interleukin‐6 (IL‐6), tumor necrosis factor receptor‐1 (TNF‐R1) and vascular endothelial growth factor as compared with control. The anti‐arthritic activity was found to be well substantiated with significant suppression of oxidative and inflammatory markers as there was decreased malonaldehyde, Nitric oxide, tumor necrosis factor alpha levels accompanied with increased glutathione and superoxide dismutase, catalase activities. Additionally, PKRE significantly inhibited the expression of degrading enzymes, matrix metalloproteinases‐3 and matrix metalloproteinases‐9 in AIA‐induced arthritic rat. Histopathology of paw tissue displayed decreased inflammatory cell infiltration as compared with control. Taken together, these results demonstrated the anti‐arthritic activity of PKRE against experimental arthritis, and the underlying mechanism behind this efficacy might be mediated by inhibition of inflammatory mediators and angiogenesis, improvement of the synovium redox status and decreased expression of matrix metalloproteinases. Copyright © 2015 John Wiley & Sons, Ltd.     

Rhodiola‐Water Extract Induces β‐endorphin Secretion to Lower Blood Pressure in Spontaneously Hypertensive Rats

Rhodiola rosea (Rhodiola) is grown at high altitudes and northern latitudes. It is mainly used clinically as an adaptogen, but antihypertensive effects have been reported for the extract. These have not been well investigated, so in the present study, we evaluated the effect of Rhodiola‐water extract on blood pressure in spontaneously hypertensive rats (SHRs) and investigated the potential mechanism(s) for this action. In conscious male SHRs, systolic blood pressure (SBP) and heart rate were recorded using the tail‐cuff method. Plasma β‐endorphin was measured by enzyme‐linked immunosorbent assay. Rhodiola‐water extract decreased SBP in SHRs in a dose‐dependent manner, and this action was more significant than that in normal group named Wistar‐Kyoto (WKY) rats. This reduction of SBP in SHRs was inhibited by pretreatment with the selective opioid μ‐receptor antagonist, cyprodime, but not by naloxonazine, an antagonist specific to opioid μ1‐receptor. Also, the SBP‐lowering action of Rhodiola‐water extract was attenuated in adrenalectomized SHRs. Moreover, Rhodiola‐water extract dose‐dependently increased β‐endorphin release in SHRs, and the elevation of β‐endorphin in SHRs was higher than that in WKY. Thus, we suggest that Rhodiola‐water extract can induce release of β‐endorphin to lower SBP in SHRs. Copyright © 2012 John Wiley & Sons, Ltd.     

In Vitro Inhibition of the Transcription Factor NF‐κB and Cyclooxygenase by Bamboo Extracts

Several bamboo species have been used in traditional medicine for the treatment of inflammatory conditions. The present study evaluates the in vitro anti‐inflammatory properties of the traditionally used bamboo species Phyllostachys nigra (Lodd.) Munro and Sasa veitchii (Carr.) Rehder to explore their future research opportunities and therapeutic potential as anti‐inflammatory agents. The extracts were evaluated for their potential inhibitory activity at the level of NF‐κB‐induced gene expression and suppression of cyclooxygenase (COX)‐1 and COX‐2 enzyme activities, representative pharmacological targets for the anti‐inflammatory action of glucocorticoids and non‐steroidal anti‐inflammatory drugs, respectively. The activity of P. nigra (Lodd.) Munro and S. veitchii (Carr.) Rehder was compared with bamboo species without traditional anti‐inflammatory indications. High‐performance liquid chromatography with diode‐array detection and liquid chromatography–tandem mass spectrometry analyses were performed to phytochemically characterize the extracts. P. nigra (Lodd.) Munro leaf extract potently inhibited NF‐κB‐induced gene expression, while S. veitchii (Carr.) Rehder leaf extract exerted a selective COX‐2 inhibition. The crude extracts consistently showed a more potent bioactivity than the solid phase extraction fractions. P. nigra (Lodd.) Munro and S. veitchii (Carr.) Rehder both exert anti‐inflammatory properties, but act via a different molecular mechanism. Copyright © 2013 John Wiley & Sons, Ltd.     

Anti‐inflammatory Activities of Gouania leptostachya Methanol Extract and its Constituent Resveratrol

Gouania leptostachya DC. var. tonkinensis Pitard. Rhamnaceae is a traditional medicinal plant used in Thailand for treating various inflammatory symptoms. However, no systematic studies have been performed concerning the anti‐inflammatory effects or molecular mechanisms of this plant. The immunopharmacological activities of a methanol extract from the leaves and twigs of G. leptostachya (Gl‐ME) were elucidated based on the gastritis symptoms of mice treated with HCl/EtOH and the inflammatory responses, such as nitric oxide (NO) release and prostaglandin E₂ (PGE₂) production, from RAW264.7 cells and peritoneal macrophages. Moreover, inhibitory target molecules were also assessed. Gl‐ME dose‐dependently diminished the secretion of NO and PGE₂ from LPS‐stimulated RAW264.7 cells and peritoneal macrophages. The gastritis lesions of HCl/EtOH‐treated mice were also attenuated after Gl‐ME treatment. The extract (50 and 300 µg/mL) clearly reduced mRNA expression of inducible NO synthase (iNOS) and cyclooxygenase (COX)‐2, nuclear translocation of p65/nuclear factor (NF)‐κB, phosphorylation of p65‐activating upstream enzymes, such as protein kinase B (AKT), inhibitor of κBα kinase (IKK), and inhibitor of κB (IκBα), and the enzymatic activity of Src. By HPLC analysis, one of the major components in the extract was revealed as resveratrol with NO and Src inhibitory activities. Moreover, this compound suppressed NO production and HCl/EtOH‐induced gastric symptoms. Therefore, these results suggest that Gl‐ME might be useful as an herbal anti‐inflammatory medicine through the inhibition of Src and NF‐κB activation pathways. The efficacy data of G. leptostachya also implies that this plant could be further tested to see whether it can be developed as potential anti‐inflammatory preparation. Copyright © 2014 John Wiley & Sons, Ltd.     

Inhibition of LPS‐Induced TNF‐α and NO Production in Mouse Macrophage and Inflammatory Response in Rat Animal Models by a Novel Ayurvedic Formulation,BV‐9238

Rheumatoid arthritis is a chronic crippling disease, where protein‐based tumor necrosis factor‐alpha (TNF‐α) inhibitors show significant relief, but with potentially fatal side effects. A need for a safe, oral, cost‐effective small molecule or phyto‐pharmaceutical is warranted. BV‐9238 is an Ayurvedic poly‐herbal formulation containing specialized standardized extracts of Withania somnifera, Boswellia serrata, Zingiber officinale and Curcuma longa. The anti‐inflammatory and anti‐arthritic effects of BV‐9238 were evaluated for inhibition of TNF‐α and nitric oxide (NO) production, in lipopolysaccharide‐stimulated, RAW 264.7, mouse macrophage cell line. BV‐9238 reduced TNF‐α and NO production, without any cytotoxic effects. Subsequently, the formulation was tested in adjuvant‐induced arthritis (AIA) and carrageenan‐induced paw edema (CPE) rat animal models. AIA was induced in rats by injecting Freund's complete adjuvant intra‐dermally in the paw, and BV‐9238 and controls were administered orally for 21 days. Arthritic scores in AIA study and inflamed paw volume in CPE study were significantly reduced upon treatment with BV‐9238. These results suggest that the anti‐inflammatory and anti‐arthritic effects of BV‐9238 are due to its inhibition of TNF‐α, and NO, and this formulation shows promise as an alternate therapy for inflammatory disorders where TNF‐α and NO play important roles. Copyright © 2014 John Wiley & Sons, Ltd.     

Inhibition of TNF‐α‐mediated endothelial cell–monocyte cell adhesion and adhesion molecules expression by the resveratrol derivative,trans‐3,5,4′‐trimethoxystilbene

Resveratrol (RSV) has been shown to have anti‐inflammatory activity and to have a protective role against atherosclerosis. Here it is shown, for the first time, that its derivative trans‐3,5,4′‐trimethoxystilbene (TMS) may be a more potent anti‐inflammatory agent than resveratrol. A comparative analysis of the inhibitory activities of related stilbenes, resveratrol, TMS and polydatin (PD), on monocyte adhesion to TNF‐α‐activated endothelial cells showed TMS to be the most effective, with PD being the least effective. RSV and its analogues inhibited, albeit differentially, the protein and mRNA expression levels of inducible cell adhesion molecules, ICAM‐1 and VCAM‐1, in cultured endothelial cells. The mechanism of the inhibitory effects of these stilbenes on endothelial cell–monocyte cell adhesion can be attributed mainly to inhibition of NF‐κB pathway activation. The results demonstrate that all three investigated stilbene compounds, especially TMS, exhibit a potent inhibitory effect on inflammation‐induced cell–cell adhesion, expression of adhesion molecules and activation of the NF‐κB pathway. Copyright © 2010 John Wiley & Sons, Ltd.     

Anti‐platelet Activity of Erythro‐(7S,8R)‐7‐acetoxy‐3,4,3′,5′‐tetramethoxy‐8‐O‐4′‐neolignan from Myristica fragrans

Platelets play a critical role in pathogenesis of cardiovascular disorders and strokes. The inhibition of platelet function is beneficial for the treatment and prevention of these diseases. In this study, we investigated the anti‐platelet activity of erythro‐(7S,8R)‐7‐acetoxy‐3,4,3′,5′‐tetramethoxy‐8‐O‐4′‐neolignan (EATN), a neolignan isolated from Myristica fragrans, using human platelets. EATN preferentially inhibited thrombin‐ and platelet‐activating factor (PAF)‐induced platelet aggregation without affecting platelet damage in a concentration‐dependent manner with IC₅₀ values of 3.2 ± 0.4 and 3.4 ± 0.3 μM, respectively. However, much higher concentrations of EATN were required to inhibit platelet aggregation induced by arachidonic acid. EATN also inhibited thrombin‐induced serotonin and ATP release, and thromboxane B₂ formation in human platelets. Moreover, EATN caused an increase in cyclic AMP (cAMP) levels and attenuated intracellular Ca²⁺ mobilization in thrombin‐activated human platelets. Therefore, we conclude that the inhibitory mechanism of EATN on platelet aggregation may increase cAMP levels and subsequently inhibit intracellular Ca²⁺ mobilization by interfering with a common signaling pathway rather than by directly inhibiting the binding of thrombin or PAF to their receptors. This is the first report of the anti‐platelet activity of EATN isolated from M. fragrans. Copyright © 2013 John Wiley & Sons, Ltd.     

Effects of Penta‐O‐galloyl‐β‐D‐glucose on Human Neutrophil Function: significant Down‐Regulation of L‐selectin Expression

Penta‐O‐galloyl‐β‐D‐glucose (PGG) occurrs in high concentrations in medicinal herbs such as Rhus chinensis, Paeonia suffruticosa, Acer truncatum and Terminalia chebula, which demonstrate anti‐inflammatory activity. We investigated the effect of PGG on stimulated and non‐stimulated neutrophils in processes which included reactive oxygen species generation (ROS), metalloproteinase‐9 and interleukin‐8 secretion (IL‐8), β₂ integrin (CD11b) and L‐selectin (CD62L) expression and apoptosis. In concentrations of 5 μM–20 μM, PGG demonstrated statistically significant inhibition of ROS generation, IL‐8 secretion and β₂ integrin expression in stimulated neutrophils. The inhibition of L‐selectin expression by PGG resulted in prevention in neutrophils’ endothelial attachment. The result obtained may explain the anti‐inflammatory activity of this compound and underline the contribution of PGG in the activity of PGG rich plant extracts. Copyright © 2012 John Wiley & Sons, Ltd.     

Methylalpinumisoflavone Inhibits Lipopolysaccharide‐Induced Inflammation in Microglial Cells by the NF‐kappaB and MAPK Signaling Pathway

Neuroinflammation is chronic inflammation within the brain that is attributed to prolonged activation of microglial cells and results in neurodegenerative events, such as neuronal dysfunction and neuronal loss. Therefore, suppression of neuroinflammation would theoretically slow progression of neurodegenerative disease. In this study, we investigated the anti‐inflammatory effects of 4′‐O‐methylalpinumisoflavone (methylalpinumisoflavone), isolated from Cudrania tricuspidata, against LPS‐induced microglial activation in BV2 cells. Exposure of BV2 cells to LPS (0.5 µg/mL) significantly increased production of pro‐inflammatory mediators, including NO, PGE₂, and pro‐inflammatory cytokines. Conversely, pre‐treatment with methylalpinumisoflavone (10 and 20 µg/mL) prior to treatment with LPS resulted in a significant decrease of LPS‐induced production of pro‐inflammatory mediators in a dose‐dependent manner. In addition, reduction of pro‐inflammatory mediators by treatment with methylalpinumisoflavone prior to treatment with LPS was accompanied by a decrease in translocation of NF‐κB p50 and p65 from the cytoplasm to the nucleus and by a decrease in activation of mitogen‐activated protein kinases (MAPKs), such as ERK1/2 and JNK. Taken together, these results suggest that methylalpinumisoflavone suppressed LPS‐induced microglial activation and production of pro‐inflammatory mediators by decreasing NF‐κB signaling and by phosphorylation of MAPKs. These results suggest the potential of methylalpinumisoflavone as an anti‐inflammatory drug candidate. Copyright © 2012 John Wiley & Sons, Ltd.     

Antiinflammatory activity and acute toxicity (LD50) of the juice of Kalanchoe brasiliensis (comb.) leaves picked before and during blooming

Kalanchoe brasiliensis Comb. (Cassulaceae) extracts from leaves picked before and during plant blooming (extracts 1 and 2, respectively) were tested for their antiinflammatory effect on carrageenin‐induced rat paw oedema and for acute toxicity (LD₅₀). Oral doses of 0.25, 0.5 and 1.0 g/kg of extract 1 significantly inhibited the paw oedema during the first 4 h after injection of 2% carrageenin, while oral doses of 0.5, 1.0 and 2.0 g/kg of extract 2 had no inhibitory activity on the paw oedema induced by carrageenin. The results indicate an antiinflammatory effect of extract 1 and a proinflammatory effect of extract 2. K. brasiliensis extracts 1 and 2 presented no acute toxicity on mice at the doses of 0.25 to 5 g/kg administered intraperitoneally. Copyright © 1999 John Wiley & Sons, Ltd.