Essential Oil from the Stem Bark of Casuarina equisetifolia Exerts Anti-inflammatory and Anti-nociceptive Activities in Rats

Abstract Herein the chemical constituents and the anti-pain properties of the essential oil from the stem bark of Casuarina equisetifolia L. (Casuarinaceae) grown in Nigeria were evaluated. The essential oil was obtained by hydrodistillation method in an all glass Clevenger-type apparatus, and characterized by gas chromatography (GC-FID) and gas chromatography-mass spectrometry (GC-MS). The hot plate method was used to determine the anti-nociceptive property whereas the anti-inflammatory activity was evaluated by carrageenan-induced and formalin experimental models. The pale-yellow essential oil was obtained in yield of 0.21% (v/w), calculated on a dry weight basis. The main constituents of the essential oil were methyl salicylate (30.4%), a-zingiberene (15.5%), (E)-anethole (9.5%), b-bisabolene (8.6%), b- sesquiphellandrene (6.9%), and ar-curcumene (6.2%). In the anti-nociceptive study, the rate of inhibition increases as the doses of essential oil increases with optimum activity at the 30th and 60th min for all tested doses. The essential oil displayed anti-nociceptive activity independently of reaction time at the highest tested dose (200 mg/kg). The essential oil of C. equisetifolia moderately reduced pain responses in early and late phases of the formalin test. The oil inhibited the paw licking in the neurogenic phase (60-63%) compared to the late phase of the formalin test. The carrageenan- induced oedema model revealed the suppression of inflammatory mediators within the 1st - 3rd h. Thus, C. equisetifolia essential oil displayed both anti-nociceptive and anti-inflammatory activities independent of the dose tested. The anti-inflammatory and anti-nociceptive activities of C. equisetifolia essential oil are herein reported for the first time

Induced volatiles in the interaction between soybean (Glycine max) and the Mexican soybean weevil (Rhyssomatus nigerrimus) / Voláteis induzidos na interação entre soja (Glycine max) e gorgulho da soja mexicana (Rhyssomatus nigerrimus)

Abstract The present study analyzed the volatile compounds emitted by Glycine max (cv. FT-Cristalina-RCH) soybean plants: healthy plants and plants damaged mechanically or by the Mexican soybean weevil Rhyssomatus nigerrimus. The SPME method was used to compare the volatile profile of soybean plants in four different conditions. The volatile profile of G. max plants infested by R. nigerrimus was qualitatively and quantitatively different from that of healthy and mechanically damaged plants. Emission of 59 compounds was detected in the four treatments. Of these compounds, 19 were identified by comparison of the Kovats index, mass spectrum and retention times with those of synthetic standards. An increase in concentration of the volatiles (Z)-3-hexenyl acetate and the compound 1-octen-3-ol was observed when the soybean plants were mechanically damaged. The compounds mostly produced by the soybean plant during infestation by male and female R. nigerrimus were 1-octen-3-ol, 6-methyl-5-hepten-2-one, (E)-β-ocimene, salicylaldehyde, unknown 10, linalool, methyl salicylate, (Z)-8-dodecenyl acetate (ester 5), ketone 2 and geranyl acetone. Behavioral effects of the identified compounds during the insect-plant interaction and their conspecifics are discussed.

Resumo O presente estudo analisou os compostos voláteis emitidos pelas plantas de soja Glycine max (cv. FT-Cristalina-RCH): plantas e plantas sadias danificadas mecanicamente ou pelo gorgulho da soja mexicana Rhyssomatus nigerrimus. O método SPME foi utilizado para comparar o perfil volátil de plantas de soja em quatro diferentes condições. O perfil volátil das plantas de G. max infestadas por R. nigerrimus foi qualitativa e quantitativamente diferente do das plantas saudáveis e danificadas mecanicamente. Foi detectada, nos quatro tratamentos, emissão de 59 compostos, dos quais 19 foram identificados por comparação do índice de Kovats, espectro de massa e tempos de retenção com aqueles de padrões sintéticos. Um aumento na concentração dos voláteis acetato de (Z)-3-hexil e do composto 1-octeno-3-ol foi observado quando as plantas de soja foram mecanicamente danificadas. Os compostos produzidos principalmente pela planta de soja durante a infestação por R. nigerrimus macho e fêmea foram 1-octeno-3-ol, 6-metil-5-hepteno-2-ona, (E)-β-ocimeno, salicilaldeído, desconhecido 10, linalol, salicilato de metila, acetato de (Z)-8-dodecenila (éster 5), cetona 2 e geranil acetona. Foram discutidos os efeitos comportamentais dos compostos identificados durante a interação inseto-planta e seus coespecíficos.

Effect of Process Change on in Vitro Release and Percutaneous Penetration of Zushima Guanjie Zhitong Gao / 中国实验方剂学杂志

Objective:To investigate the effect of the production process of Zushima Guanjie Zhitong Gao from solvent method to hot-pressed method on <italic>in vitro</italic> kinetic behavior of this preparation. Method:Solvent and hot-pressed methods were used to prepare three batches of samples above pilot scale， and <italic>in vitro</italic> release and percutaneous penetration of the index components （7，8-dihydroxycoumarin and methyl salicylate） in Zushima Guanjie Zhitong Gao were investigated by modified Franz diffusing cells. Result:The contents of 7，8-dihydroxycoumarin and methyl salicylate in Zushima Guanjie Zhitong Gao prepared by solvent method were 73.72， 494.67 μg/patch， and their contents in hot-pressed method samples were 159.21， 2 638.99 μg/patch， respectively. In the solvent method samples， the average cumulative release amounts of 7，8-dihydroxycoumarin and methyl salicylate in 24 h were 2.04， 12.21 μg， and their average cumulative release amounts in 24 h of hot-pressed method samples were 2.16， 36.24 μg， respectively. In the solvent method samples， the average cumulative permeation amounts of 7，8-dihydroxycoumarin and methyl salicylate in 24 h were 0.38， 2.79 μg， and they were 0.40， 7.49 μg in hot-pressed method samples. The cumulative release and permeation amounts in 24 h of 7，8-dihydroxycoumarin in the hot-pressed method samples were basically the same as those of the solvent method samples， but the cumulative release and permeation amounts in 24 h of methyl salicylate in the hot-pressed method samples were significantly higher than those of the solvent method samples （<italic>P</italic><0.05）. Conclusion:The retention of 7，8-dihydroxycoumarin and methyl salicylate by hot-pressed method is better than that of the solvent method. The process change has no significant effect on the <italic>in vitro</italic> kinetics of 7，8-dihydroxycoumarin in Zushima Guanjie Zhitong Gao， however， after the change from the solvent method to the hot-pressed method， the methyl salicylate in this preparation has a higher cumulative release and permeation amounts.

Desarrollo y validación de un método de cromatografía gaseosa para la determinación de mentol, salicilato de metilo, timol y ácido benzoico en solución antiséptica / Development and validation of a gas chromatographic method for the determination of menthol, methyl salicylate, thymol, and benzoic acid in antiseptic solution

RESUMEN Se desarrolló y validó un método por cromatografía gaseosa para la cuantificación simultánea de mentol (MT), salicilato de metilo (SM), timol (TM) y ácido benzoico (AB) en una solución antiséptica bucal. Se utilizó una columna DB-WAX y dietilenglicol como estándar interno. Se determinó la linealidad en un intervalo de concentraciones de 64,00 a 96,00 µg/mL (MT), 80,00 a 120,00 µg/mL (SM), 96,00 a 144,00 µg/mL (TM) y 48,00 a 72,00 µg/mL (AB), obteniendo coeficientes de correlación superiores a 0,999, y coeficientes de variación de los factores de respuestas de 1,18, 1,95, 3,52 y 1,48%, respectivamente. Se establecieron límites de detección de 0,51; 1,14; 3,34 y 1,402 ng/mL para el MT, SM, TM y AB, respectivamente, mientras los límites de cuantificación fueron de 1,45, 3, 43, 9, 73 y 4, 36 ng/mL en cada caso. Los porcentajes de recuperación fueron de 100,03, 99, 31, 99, 92 y 100,12; con coeficientes de variación de 0,42, 0,79, 0,66 y 0,76% para cada caso. El método fue lineal, exacto, preciso y selectivo para la determinación de los analitos en el control de calidad.

SUMMARY A method was developed and validated by gas chromatography for the simultaneous quantification of menthol (MT), methyl salicylate (SM), thymol (TM) and benzoic acid (AB) in an oral antiseptic solution. A DB-WAX column and diethylene glycol was used as internal standard. Linearity was determined in a concentration range of 64.00 to 96.00 µg/mL (MT), 80.00 to 120.00 µg/mL (SM), 96.00 to 144.00 µg/mL (TM) and 48.00 to 72.00 µg/mL (AB) achieving correlation coefficients greater than 0.999, and coefficients of variation of the response factors of 1.18, 1.95, 3.52 and 1.48%, respectively. Detection limits were established: 0.51, 1.14, 3.34 and 1.402 ng/mL for MT, SM, TM, and AB, respectively, while the quantification limits were 1.45, 3.43, 9.73 and 4.36 ng/mL in each case. Recovery percentages were 100.03, 99.31, 99.92 and 100.12; with coefficients of variation of 0.42, 0.79, 0.66 and 0.76% for each case. The method was linear, accurate, precise, and selective for the determination of analytes in quality control.

Chemical constituents from anti-inflammatory and analgesic active fraction of Gaultheria leucocarpa var. yunnanensis / 中草药

Objective To study the chemical constituents from anti-inflammatory and analgesic active fraction (ARF) of Gaultheria leucocarpa var. yunnanensis. Methods The compounds were isolated and purified by various techniques of column chromatography, and their structures were determined according to physicochemical properties and spectral analyses. Results Sixteen compounds were obtained and identified as methyl salicylate 2-O-β-D-glucopyranoside (1), gaultherin (2), MSTG-A (3), MSTG-B (4), ethyl-O-β-D-xylopyranoside (5), ethyl-O-β-D-xylopyranosyl (1→6)-O-β-D-glucopyranoside (6), methyl-O-β-D-xylopyranosyl (1→6)-O-β-D-glucopyranoside (7), roseoside (8), paeoniflorin (9), vanillic acid (10), 2,5-dihydroxybenzoic acid (11), 3,4-dimethoxycinnamic acid (12), ferulic acid (13), chlorogenic acid (14), 4-hydroxy-2,6-dimethoxyphenol-O-β-D-glucopyranoside (15), and 3-methoxyl-1H-pyrrole (16). Conclusion Compounds 5-9, 12, 15, and 16 are obtained from this plant and compound 7 is isolated from the plants of genus Gaultheria Kalm ex L. for the first time. Compounds 8, 9, 12, and 15 are obtained from family Ericaceae for the first time.

Induced volatiles in the interaction between soybean (Glycine max) and the Mexican soybean weevil (Rhyssomatus nigerrimus)

Abstract The present study analyzed the volatile compounds emitted by Glycine max (cv. FT-Cristalina-RCH) soybean plants: healthy plants and plants damaged mechanically or by the Mexican soybean weevil Rhyssomatus nigerrimus. The SPME method was used to compare the volatile profile of soybean plants in four different conditions. The volatile profile of G. max plants infested by R. nigerrimus was qualitatively and quantitatively different from that of healthy and mechanically damaged plants. Emission of 59 compounds was detected in the four treatments. Of these compounds, 19 were identified by comparison of the Kovats index, mass spectrum and retention times with those of synthetic standards. An increase in concentration of the volatiles (Z)-3-hexenyl acetate and the compound 1-octen-3-ol was observed when the soybean plants were mechanically damaged. The compounds mostly produced by the soybean plant during infestation by male and female R. nigerrimus were 1-octen-3-ol, 6-methyl-5-hepten-2-one, (E)--ocimene, salicylaldehyde, unknown 10, linalool, methyl salicylate, (Z)-8-dodecenyl acetate (ester 5), ketone 2 and geranyl acetone. Behavioral effects of the identified compounds during the insect-plant interaction and their conspecifics are discussed.

Resumo O presente estudo analisou os compostos voláteis emitidos pelas plantas de soja Glycine max (cv. FT-Cristalina-RCH): plantas e plantas sadias danificadas mecanicamente ou pelo gorgulho da soja mexicana Rhyssomatus nigerrimus. O método SPME foi utilizado para comparar o perfil volátil de plantas de soja em quatro diferentes condições. O perfil volátil das plantas de G. max infestadas por R. nigerrimus foi qualitativa e quantitativamente diferente do das plantas saudáveis e danificadas mecanicamente. Foi detectada, nos quatro tratamentos, emissão de 59 compostos, dos quais 19 foram identificados por comparação do índice de Kovats, espectro de massa e tempos de retenção com aqueles de padrões sintéticos. Um aumento na concentração dos voláteis acetato de (Z)-3-hexil e do composto 1-octeno-3-ol foi observado quando as plantas de soja foram mecanicamente danificadas. Os compostos produzidos principalmente pela planta de soja durante a infestação por R. nigerrimus macho e fêmea foram 1-octeno-3-ol, 6-metil-5-hepteno-2-ona, (E)--ocimeno, salicilaldeído, desconhecido 10, linalol, salicilato de metila, acetato de (Z)-8-dodecenila (éster 5), cetona 2 e geranil acetona. Foram discutidos os efeitos comportamentais dos compostos identificados durante a interação inseto-planta e seus coespecíficos.

Study on Quality Standard of Miao Medicine Gaultheria yunnanensis / 中国药房

OBJECTIVE:To establish the quality standard for Gaultheria yunnanensis. METHODS:TLC was adopted for quali-tative identification of samples. Moisture,total ash and acid-insoluble ash were determined. HPLC method was used to determine the content of methyl salicylate. The determination was performed on Eclipse XDB-C18 column with mobile phase consisted of meth-anol-water(62:38,V/V)at the flow rate of 1.0 mL/min. The detection wavelength was set at 307 nm,and column temperature was 30 ℃. The sample size was 10 μL. RESULTS:TLC spots of samples were clear and well separated. The moisture was 8.2%-10.8%,total ash was 0.9%-4.0% and acid-insoluble ash was 0.1%-0.9%. The linear range of methyl salicylate were 0.045-0.73μg(r=0.9999). RSDs of precision,stability and reproducibility tests were no more than 1.0%. The recoveries of meth-yl salicylate were 97.8%-104.3%(RSD=2.6%,n=9). CONCLUSIONS:The established standard can be used for quality control of G. yunnanensis.

The Vasodilator Effect of a Cream Containing 10% Menthol and 15% Methyl Salicylate on Random-Pattern Skin Flaps in Rats

BACKGROUND: It is still difficult to prevent partial or full-thickness flap necrosis. In this study, the effects of a cream containing menthol and methyl salicylate on the viability of randompattern skin flaps were studied. METHODS: Forty female Sprague-Dawley rats were divided into two equal groups. Caudally based dorsal random-pattern skin flaps were elevated, including the panniculus carnosus. In the study group, 1.5 mL of a cream containing menthol and methyl salicylate was applied to the skin of the flap, and saline solution (0.9%) was used in the control group. Upon completion of the experiment, flap necrosis was analyzed with imaging software and radionuclide scintigraphy. Histopathological measurements were made of the percentage of viable flaps, the number of vessels, and the width of the panniculus carnosus muscle. RESULTS: According to the photographic analysis, the mean viable flap surface area in the study group was larger than that in the control group (P=0.004). According to the scintigrams, no change in radioactivity uptake was seen in the study group (P>0.05). However, a significant decrease was observed in the control group (P=0.006). No statistically significant differences were observed between the groups in terms of the percentage of viable flaps, the number of vessels, or the width of the panniculus carnosus muscle (P>0.05). CONCLUSIONS: Based on these results, it is certain that the cream did not reduce the viability of the flaps. Due to its vasodilatory effect, it can be used as a component of the dressing in reconstructive operations where skin perfusion is compromised.

Development of colorimetric method for the quantification of methyl salicylate in bulk and formulations.

Methyl salicylate is chemically methyl-O-hydroxybenzoate and act as an active ingredient in many topical analgesic preparations. It posses good counter irritant activity. In this present study, a colorimetric method is developed for the quantitative estimation of methyl salicylate, both in the bulk form and in the formulations. A dark bluish – purple colour is formed between methyl salicylate and ferric chloride reagent. The intensity of the colour is directly related to the concentration of methyl salicylate present. The developed method was simple and gave accurate and precise results. The developed colour had its maximum value of absorbance at 537 nm when scanned in the visible region. The absorbance of the coloured system was measured at 537 nm. Beer’s law was obeyed in the range of 12 to 72 μg/mL. The equation for the straight line was found to be, y = 0.0075 x + 0.002 with correlation coefficient value of 0.996. The developed method was applied to semi – solid formulations for the quantitative estimation of methyl salicylate. The percentage of recovery was between 99.62 % in the formulation. The method proved to be a convenient analytical tool for the quantity determination of methyl salicylate.

Composição e variação sazonal do óleo essencial de Myrcia obtecta (O. Berg) Kiaersk. var. obtecta, Myrtaceae / Composition and seasonal variation of essential oils of Myrcia obtecta (O. Berg) Kiaersk. var. obtecta, Myrtaceae

Os óleos essenciais, isolados por hidrodestilação de folhas e flores de Myrcia obtecta (O. Berg) Kiaersk. var. obtecta, Myrtaceae, coletadas a cada dois meses em Curitiba, Paraná, durante o período de outubro/2005 a agosto/2006, foram analisados por CG e CG/EM. Os óleos das folhas foram caracterizados pelo elevado conteúdo de sesquiterpenos cíclicos (cerca de 70 por cento) e quantidades variáveis de monoterpenos (5,6-16,7 por cento), compostos alifáticos (0,4-5,5 por cento) e salicilato de metila (0,1-1,5 por cento). Foram identificados 55 componentes, sendo que trans-calameneno (17,029,3 por cento) foi o componente majoritário em todas as amostras. Não houve mudanças significativas na composição, excepto em outubro (floração) quando α-terpineol e trans-calameneno atingiram os maiores níveis (11,2 por cento e 29,3 por cento, respectivamente). O óleo das flores contém salicilato de metila (89 por cento) como constituinte majoritário.

The essential oils isolated by hydrodistillation from leaves and flowers of Myrcia obtecta (O. Berg) Kiaersk. var. obtecta, Myrtaceae, collected bimonthly in Curitiba, Paraná state, Brazil, from October/2005 to August/2006 were analyzed by capillary GC and GC/MS. Fifty-five components were identified in the leaf oils, which were characterized by predominance of cyclic sesquiterpenes (around 70 percent) and variable amounts of monoterpenes (5.616.7 percent), aliphatic compounds (0.4-5.5 percent) and methyl salicylate (0.1-1.5 percent). The major component in all samples was trans-calamenene (17.0-29.3 percent). The composition did not show significant seasonal variation, except in October (flowering) when α-terpineol and trans-calamenene peaked (11.2 percent and 29.3 percent, respectively). The flower oil was characterized by high content of methyl salicylate (89 percent).

Determination of Menthol and Methyl Salicylate in Shangtongning Adhesive Plaster by Capillary Gas Chromatography / 中国中医药信息杂志

Objective To establish a capillary gas chromatography method for the determination of menthol and methyl salicylate in Shangtongning adhesive plaster. Methods The internal standard substance of n-Pentadecane was employed. The GC system consisted of a DB-WAX capillary column (30 m?0.32 mm, 0.25 ?m), nitrogen as the carrier gas, column temperature at 130 ℃, FID as the detector at 200 ℃, and with a split ratio of 12∶1. Results The contents of menthol and methyl salicylate in 6 lots of samples were determined and the constituents concerned were separated well under the chromatographic condition. The average recovery of menthol and methyl salicylate were 101.52% (RSD=2.66%) and 100.34% (RSD=3.38%), respectively. Conclusion The method was sensitive, accurate, specific and suitable for the quality control of the preparation.

Determination of four volatiles in Baicao Oil by GC with large bore capillary column / 中成药

AIM:To establish a method for determining menthol, methyl salicylate,cinnamal dehyde and engenol simultaneously in Baicao Oil by GC. METHODS: A separation was performed on a large bore capillary column(30m?0.53mm) with PEG-20M as the stationary phase of GC.The method was carried out with FID as the detector without split stream sampling.A programmed temperature was employed.Benzyl alcohol was selected as the internal standard substance. RESULTS: An excellent seperation of menthol,methyl salicylate,cinnamal and engenol in Baicao Oil was gained.The good linear relationships of menthol (62.7?10~3-2.01?10~3 ?g/mL),methyl salicylate (119?10~3-3.81?10~3 ?g/mL),cinnamal dehyde (0.42-205 ?g/mL),engenol (19.9-636 ?g/mL) were observed.All correlation coefficients were greater than (0.999 2).The average recovery of menthol,methyl salicylate,cinnamal and engenol were between(98.6%)～100.3%,and RSD was less than 2.4%. CONCLUSION: The method is simple,accurate and suitable for quality control of Baicao Oil.

Simultaneous determination of camphor,menthol,synthetic borneol and methyl salicylate in Shexiang Zhuanggu Plaster by GC / 中成药

AIM: To establish the simultaneous determination of camphor,menthol,synthetic borneol and methyl salicylate in Shexiang Zhuanggu Plaster(Camphor,Mentholum,Borneolum Syntheticum,etc). by GC. METHODS: The sample solution was distilled in volatile oil determination apparatus.Naphthalene was used for the internal standard.The GC system consisted of capillary column,PEG-20M as the stationary phase,nitrogen as the carrier gas,FID as the detector.The programmed temperature-GC and internal standard method was employed to determine four kinds of components in Shexiang Zhuanggu Plaster. RESULTS: Camphor,menthol,synthetic borneol(borneol and isoborneol) and methyl salicylate in Shexiang Zhuanggu Plaster and naphthalene have been separated well under the same chromatographic condition.The average recovery of camphor,menthol,synthetic borneol and methyl salicylate were 96.92%(RSD=2.42%),98.03%(RSD=1.81%),99.02%(RSD=1.47%) and 98.15%(RSD=1.59%),respectively. CONCLUSION: The method is simple,accurate and separable,and can be used to control the quality of Shexiang Zhuanggu Plaster

Determination of camphor, menthol, synthetic borneol and methyl salicylate in four kinds of TCM adhesive plasters by GC / 中成药

AIM: To establish the determination for camphor, menthol, synthetic borneol, and methyl salicylate in four kinds of TCM adhesive plasters by GC. METHODS: The internal standard method was employed. Naphthalene, the internal standard, was added to the sample before treatment. The sample solution was prepared by distillation with essential oil collector. The GC system consisted of glass column, 10% PEG 20M as the stationary phase, nitrogen as the carrier gas, column temperature at 130℃, and FID as the detector. RESULTS: Camphor, menthol, synthetic borneol(borneol and isoborneol), and methyl salicylate in 6 lots of samples and naphthalene have been separated well under the chromatographic condition. In the methodology study on Fengshi Zhitong Adhesive Plasters, the average recovery of camphor, menthol, synthetic borneol, and methyl salicylate were 95.54%( RSD =1.6%), 96.90%( RSD =2.2%), 97.30%( RSD =1.4%), and 96.48%( RSD =2.8%), respectively. CONCLUSION: The method is sensitive, accurate and reproducible, and can be used to control the quality of the four kinds of preparations.