Systematic identification of chemical forms of key terpene synthase in Cinnamomum camphora / 中国中药杂志

Cinnamomum camphora is an important economic tree species in China. According to the type and content of main components in the volatile oil of leaf, C. camphora were divided into five chemotypes, including borneol-type, camphor-type, linalool-type, cineole-type, and nerolidol-type. Terpene synthase(TPS) is the key enzyme for the formation of these compounds. Although several key enzyme genes have been identified, the biosynthetic pathway of(+)-borneol, which has the most economic value, has not been reported. In this study, nine terpenoid synthase genes CcTPS1-CcTPS9 were cloned through transcriptome analysis of four chemical-type leaves. After the recombinant protein was induced by Escherichia coli, geranyl pyrophosphate(GPP) and farnesyl pyrophosphate(FPP) were used as substrates for enzymatic reaction, respectively. Both CcTPS1 and CcTPS9 could catalyze GPP to produce bornyl pyrophosphate, which could be hydrolyzed by phosphohydrolase to obtain(+)-borneol, and the product of(+)-borneol accounted for 0.4% and 89.3%, respectively. Both CcTPS3 and CcTPS6 could catalyze GPP to generate a single product linalool, and CcTPS6 could also react with FPP to generate nerolidol. CcTPS8 reacted with GPP to produce 1,8-cineol(30.71%). Nine terpene synthases produced 9 monoterpene and 6 sesquiterpenes. The study has identified the key enzyme genes responsible for borneol biosynthesis in C. camphora for the first time, laying a foundation for further elucidating the molecular mechanism of chemical type formation and cultivating new varieties of borneol with high yield by using bioengineering technology.

Chemical constituents from the leaves of Cinnamomum camphora var. linaloolifera and their anti-inflammatory activities / 中国中药杂志

Thirteen compounds were isolated and purified from the leaves of Cinnamomum camphora by the macroporous resin,silica gel,and Sephadex LH-20 column chromatographies. Those compounds were further identified by IR,UV,MS,and NMR techniques:( 2 S)-1-( 3″,4″-methylenedioxy phenyl)-3-( 2',6'-dimethoxy-4'-hydroxyphenyl)-propan-2-ol( 1),( 2 R,3 R)-5,7-dimethoxy-3',4'-methylenedioxy flavanol( 2),9-hydroxysesamin( 3),sesamin( 4),piperitol( 5),kobusin( 6),(-)-aptosimon( 7),acuminatolide( 8),1β,11-dihydroxy-5-eudesmene( 9),lasiodiplodin( 10),vanillin( 11),p-hydroxybenzaldehyde( 12),and p-hydroxybenzoic acid ethyl ester( 13). Compound 1 was a novel compound,and compounds 2,6,7,9 and 10 were isolated from Cinnamomum plants for the first time. Compounds 4,7 and 10 were found to possess good inhibitory effect on IL-6 production in LPS-induced BV2 cells at a concentration of 20 μmol·L-1 in the in vitro bioassay,with inhibition rates of 51. 26% ± 4. 13%,67. 82% ± 3. 77% and85. 81%±1. 19%,respectively.

Cloning and expression analysis of 5-phosphomevalonate kinase gene (CcPMK) in Cinnamomum camphora / 中国中药杂志

The 5-phosphomevalonate kinase(PMK) is a key enzyme in mevalonate(MVA) pathway which reversibly catalyzes the phosphorylation of mevalonate 5-phosphate(MVAP) to form mevalonate-5-diphosphate(MVAPP) in the presence of ATP and divalent metal ion such as Mg~(2+). In this research, on the basis of the transciptome database of Cinnamomum camphora, the PMK was cloned by cDNA from C. camphora, and was named CcPMK(GenBank number KU886266). The ORF of CcPMK was composed of 1 545 bp, encoding 514 amino acids. The bioinformatics analysis of CcPMK indicated that the molecular weight of the encoded protein was 56.14 kDa, with a theoretically isoelectric point of 7.64, and there was no signal peptide and transmembrane structure in putative protein. By multiple sequence alignment and phylogenetic tree analysis, we found that similarity between CcPMK and PMK amino acid sequence of other plants was as high as 75%. Among the similar sequences, 45% of them belonged to the alpha helix, while 16% belonged to the beta strand. CcPMK obtained 3 PMK protein family motifs and 1 ATP binding site Gly-Leu-Gly-Ser-Ser-Ala-Ala, and its 3 D structure contained a catalytic pocket structure, proving CcPMK as a member of PMK gene family. The result of phylogenetic tree showed that CcPMK was closely related to monocotyledon plants such as Phonenix dactylifera. The results of the Real-time PCR indicated that the expression level of CcPMK in borneol type was higher than that in linalool type, cineol type, iso-nerolidol type and camphor type. CcPMK expressed highest in roots and lowest in branches. Our results revealed that the expression level of CcPMK was different among five chemical types and different plant tissues, and the research provides foundation for further study of the terpenoids biosynthetic pathway in C. camphora.

Chemical composition and larvicidal activities of essential oil of Cinnamomum camphora (L. ) leaf against Anopheles stephensi

Abstract INTRODUCTION Anopheles stephensi is the main malaria vector in Southeast Asia. Recently, plant-sourced larvicides are attracting great interests. METHODS: The essential oil was extracted from the leaf of Cinnamomum camphora (L.), and a bioassay was conducted to determine the larvicidal efficacy. The chemical composition of the essential oil was determined by GC-MS analysis. RESULTS: The oil showed strong, dose-dependent larvicidal activities. The onset of larvicidal efficiency was rapid. The LC50 and LC95 were determined as 0.146% and 1.057% at 1 h, 0.031% and 0.237% at 12 h, 0.026% and 0.128% at 24 h, respectively. The oil contains 32 compounds. CONCLUSIONS The essential oil of C. camphora leaf has an excellent larvicidal potential for the control of A. stephensi.

Adaptação homóloga e heteróloga de células sésseis de Cronobacter sakazakii a óleos essenciais / Homologous and heterologous adaptation of Cronobacter sakazakii sessile cells to essential oils

Cronobacter sakazakii é patogênico, importante contaminante de fórmulas infantis, que apresenta tolerância aos sanificantes comerciais. Assim, os óleos essenciais se destacam. O objetivo desse trabalho foi avaliar a adaptação homologa e heteróloga de células sésseis de C. sakazakii aos óleos de canela, ho wood e menta. C. sakazakii foi adaptado a concentrações subletais dos óleos e desafiado a diferentes concentrações dos mesmos, determinando-se a adaptação homóloga. Já para avaliação da adaptação heteróloga os óleos foram diferentes daqueles usados na adaptação. De modo geral a adaptação tornou as células sésseis mais sensíveis aos óleos, entretanto, houve adaptação homóloga para o óleo de canela. Também foi observada adaptação heteróloga. C. sakazakii na forma séssil foi capaz de desenvolver adaptação homóloga e heteróloga.

Insecticidal and repellent activity of the essential oil of Cinnamomum camphora var. linaloolifera Y. Fujita (Ho-Sho) and Cinnamomum camphora (L. ) J Presl. var. hosyo (Hon-Sho) on Sitophilus zeamaisMots. (Coleoptera, Curculionedae) / Atividade inseticida e repelente do óleo essencial de Cinnamomum camphora var. linalooliferaY. Fujita (Ho-Sho) e Cinnamomum camphora (L. ) J Presl. var. hosyo (Hon-Sho) sobre Sitophilus zeamais Mots. (Coleoptera, Curculionedae).

ABSTRACT: The aim of this work was to evaluate the insecticidal and repellency activity of the essential oil of Cinnamomum camphora var. linaloolifera Y. Fujita (Ho-Sho) and Cinnamomum camphora (L.) J Presl.var. hosyo (Hon-Sho), against the Sitophilus zeamais in maize grains. The essential oils were obtained by hydrodistillation and analyzed by GC-MS.The insecticidal activity was determined by the toxicity of different concentrations of essential oils during 24 hours of contact with the insects, in the absence of feed substrate. The Bioassays of repellency were conducted with lethal doses (LD50,LD25,and LD12.5) obtained from insecticidal bioassay. In order to compare the treatments the preference index (PI) was employed. The analysis of the essential oils of Cinnamomum camphora leaves indicated 68% of camphor and 9% of linalool for the variation Hon-Sho and 95% of linalool to the variation Ho-Sho. The variation Ho-Sho presented greatest insecticidal activity than the variation Hon-Sho against the Sitophilus zeamais, with LD50 of 0.35 µL/cm2, whereas in the variation Hon-Sho the ratewas 0.48 µL/cm2. However, considering only the concentrations of linalool and camphor of Ho-Sho and Hon-Sho, the lethal doses' evaluation of these compounds were similar. The values of the preference index ranged from -0.3 to -0.8 for thevariation Ho-Sho and -0.2 to -0.7 for the variation Hon-Sho. The essential oils evaluated in this work showed repellent activity against Sitophiluszeamais in vitro and in trials performed in mini-silos

RESUMO: Atividade inseticida e repelente do óleo essencial de Cinnamomum camphora var. linaloolifera Y. Fujita (Ho-Sho) e Cinnamomum camphora (L.) J Presl. var. hosyo (Hon-Sho) sobre Sitophilus zeamais Mots. (Coleoptera, Curculionedae). O objetivo deste trabalho foi avaliar a atividade inseticida e de repelência dos óleos essenciais de Cinnamomum camphora var. linaloolifera Y. Fujita (Ho-Sho) e Cinnamomum camphora (L.) J Presl. var. hosyo (Hon-Sho) contra Sitophilus zeamais em grãos de milho. Os óleos essenciais foram obtidos por hidrodestilação e analisados por CG-EM. A atividade inseticida foi determinada pela toxicidade de diferentes concentrações dos óleos essenciais durante 24 horas de contato com os insetos, na ausência de substrato alimentar. Os bioensaios de repelência foram realizados com as doses letais (DL50, DL25, e DL12,5) obtidas do bioensaio inseticida. Para comparar os tratamentos foi utilizado o índice de preferência (PI). A análise de óleos essenciais de folhas de Cinnamomum camphora indicou a presença de 68% de cânfora e 9% de linalol na var. Hon-Sho e de 95% de linalol na var. Ho-Sho. A var. Ho-Sho apresentou maior toxicidade que var. Hon-Sho contra Sitophilus zeamais, com DL50 de 0,35 µL/cm2,enquanto que na var. Hon-Sho foi de 0,48 µL/cm2. No entanto, considerando apenas as concentrações de linalol e cânfora de Ho-Sho e Hon-Sho, a avaliação das doses letais destes compostos foram semelhantes. Os valores do índice de preferência variaram de -0,3 a -0,8 para a var. Ho-Sho e -0,2 para -0,7 para a var. Hon-Sho. Os óleos essenciais avaliados neste trabalho apresentaram atividade repelente contra Sitophilus zeamais in vitro e em experimentos realizados em mini-silos

Screening the antifungal activity of essential oils against decay fungi from palmyrah leaf handicrafts

BACKGROUND: The whitish tender leaves of Palmyrah are used for making handicrafts. The problem with these articles is discolouration with time and become more brittle due to fungal attack. This could be prevented by some protective coating. Instead of expensive and harmful chemicals we decided to test natural plant essential oils to control fungal attack. Palmyrah leaf article decay fungi were isolated from two different sites of Jaffna peninsula. In this investigation Antifungal Activity of different plant essential oils from neem (Azadirachta indica), castor (Ricinus communis), citronella (Cymbopogon sp) and camphor (Cinnamomum camphora) obtained from local market have been evaluated against isolated fungi. For screening of Antifungal activity, tests and controls were set to determine minimum inhibitory concentration (MIC) and Percentage of Growth Inhibition. RESULTS: Morphologically three different types of Palmyrah leaf decay fungi were isolated and characterized asAspergillus niger, Aspergillus flavus and Penicillium sp. Neem and castor oils have recorded no significant (0.05 > P) antifungal activity while citronella and camphor oils showed significantly different antifungal activity compared with control. Camphor oil and Citronella oil showed 100, 58.13% of average growth inhibition for A. niger. 96.38, 51.32% for A.flavus and 84.99, 72.76% forPenicillium sp respectively. Camphor oil showed highest percentage of growth inhibition at lowest minimum inhibitory concentration compared with citronella oil. Camphor oil was found to be highly antifungal and most effective against A niger, and A. flavus, compared with Penicillium sp and gave 100 percentage of growth inhibitions at 5, 1 and 15 ml/dl minimum inhibitory concentration respectively. CONCLUSION: Significantly higher broad-spectrum of antifungal activity was observed in camphor oil than other tested oils because it showed highest percentage of growth inhibition at lowest inhibitory concentration. Therefore it could be used for the development of new environmental friendly antifungal agent for the preservation of leafy handicrafts. Further formulation, field experiments are necessary to achieve this target.

Volatile metabolites analysis and molecular identification of endophytic fungi bn12 from Cinnamomum camphora chvar. borneol / 中国中药杂志

<p><b>OBJECTIVE</b>To identify endophytic fungi bn12 from Cinnamomum camphora chvar, borneol and analysis its volatile metabolites.</p><p><b>METHOD</b>The endophytic fungi bn12 was identified by morphological observation. volatile metabolites of endophytic fungi bn12 was analyzed by gas chromatography/mass spectrography (GC-MS).</p><p><b>RESULT</b>Volatile metabolites of endophytic fungi bn12 contain borneol and much indoles. The ITS sequence of endophytic fungi bnl2 is most similar to the ITS sequence of pleosporaceae fungus, particularly C. nisikadoi.</p><p><b>CONCLUSION</b>Endophytic fungi bn12 is belong to pleosporaceae fungus. It has the ability of producing broneol.</p>

Callus induction of Cinnamonum camphora and formation of borneol / 中国中药杂志

<p><b>OBJECTIVE</b>To optimize the condition of callus of Cinnamonum camphora induced.</p><p><b>METHOD</b>GC and plant tissue culture method were applied in the study.</p><p><b>RESULT</b>The effect of callus induced and the growth of callus were different in MS medium with different proportion of hormone. The ration of callus induced was the highest and the growth of callus was the most prosperous in the MS medium with 4 mg x L(-1) 2,4-D and 0.2 mg x L(-1) 6-BA. It is found that callus induced by young leaf contained borneol, but callus induced by young stem not.</p><p><b>CONCLUSION</b>The optimization of callus of C. camphora induced is using the MS medium with 4 mg x L(-1) 2,4-D + 0.2 mg x L(-1) 6-BA. Callus induced by young leaf can generate borneol.</p>

Determination of d-borneol in the different parts of Cinnamomum camphora by GC-MS / 中国中药杂志

<p><b>OBJECTIVE</b>To measure and compare the content of d-borneol in the different parts of Cinnamomum camphora by GC-MS.</p><p><b>METHOD</b>With water-steam distillation and GC-MS method, d-borneol was extracted and determined.</p><p><b>RESULT</b>The linear range of d-borneol was 0.4-2.8 microg (r = 0.999 9). The average recovery was 95.40%, and RSD was 0.56%.</p><p><b>CONCLUSION</b>The method is simple and accurate with good separation. The content of d-borneol in the dried leaves 63.97% in the crude exfract. It can provide the proof of the exploitation of C. camphora.</p>

Study on antiinflammatory effect of different chemotype of Cinnamomum camphora on rat arthritis model induced by Freund's adjuvant / 中国中药杂志

<p><b>OBJECTIVE</b>To study the antiinflammatory effects of naphtha from different chemotypes of Cinnamomum camphora and natural borneol on the rat arthritis model induced by Freund's adjuvant.</p><p><b>METHOD</b>The arthritis model was induced by injecting Freund's adjuvant in rat voix pedis dermis and the rats were randomly divided into seven groups: normal control group, model control group, triptergium wilfordii control group, borneol chemotype naphtha group, camphor chemotype naphtha group, isocamphane chemotype naphtha group and natural borneol group. Rats of the triptergium wilfordii control group were given orally 8.1 mg x kg(-1) triptergium wilfordii for 35 days, rats of the normal control group and model control group were given same volume water, and rats of other groups were given 80 mg x kg(-1) corresponding drug. We observed the rat common condition, weighed the rat body weight weekly, measured the degree of swelling of voix pedis every 4 days, weighed the thymus and spleen on the end of life, and measured the contents of cell factor TNF-alpha, IL-2, and IL-6 in rat blood serum.</p><p><b>RESULT</b>As far as the arthrosis degree of swelling and the contents of cell factor TNF-alpha, IL-2, IL-6 were concerned, rats of model control group were higher than normal control group, and rats of other drug groups were lower than the model control group. The order of inhibition ratios of the arthrosis degree of swelling from high to low principle was isocamphane chemotype naphtha group, camphor chemotype naphtha group, borneol chemotype naphtha group and natural borneol group. All medication administration teams evidently reduced the contents of the IL-2 and IL-6, and the inhibition ratios were higher than 38%. In the case of the contents of TNF-alpha and IL-2, all groups were not evidently different. In the case of inhibition of IL-6, camphor chemotype naphtha group was better than borneol chemotype naphtha group and natural borneol group, the latter was better than isocamphane chemotype naphtha group. As far as the weight, thymus index and spleen index were concerned, all medication administration groups were not different.</p><p><b>CONCLUSION</b>The different chemotypes of C. camphora have anti-inflammatory effect on the rat arthritis model induced by Freund's adjuvant, but pharmacological activity and mechanism of action are different. The study points out the clinical curative effects of the chemotypes of the kindred medicinal plant are different, and please consider the difference of chemotype in clinical application.</p>

Evaluation of oils from the plants mentha sativa,] ocimum basilicum and cinnamomum camphor on survival, hepatic enzymes activities and protein electrophoresis of biomphalaria Alexandrina snails

Three plant oils from the plants Spearmint, Basil and Camphor were tested for their molluscicidal activity against Biomaphalaria alexandrina snails. Spearmint oil was more toxic than the other two oils against B. alexandrina. Its Lc50 was 42 ppm; meanwhile the values were 500 and 560 ppm for Basil and Camphor, respectively. Spearmint oil, also, showed a more deleterious effect against both miracidia and cercariae of S. mansoni than the other two oils, all of them were killed after 15 min of exposure to 70 ppm. The present results indicated that the alterations in total protein concentrations in tissues of treated snails were not significantly different from that of control ones. It was noticed, also, that the increase in the activity of ALT enzyme and the decrease in that of AlkP enzyme in tissues of treated snails were not significantly different than that of control snails. However, Spearmint oil at 32 and 42 ppm significantly decreased the activities of AST and ACP activities in snail's tissues in comparison with control ones. The electrophoretic [SDS-PAGE] pattern of tissue soluble proteins extracted from treated and controls B. alexandrina snails showed different plant oils effects on the synthesis of protein within snails yielded a complex pattern of polypeptides ranging in molecular weight between 14.7 to 304.6 KDa. Some bands were present in treated snails especially at Camphor oil and at 500ppm of Basil oil. At least, three bands were absent from tissues of snails treated with 42ppm of Spearmint oil in comparison with control group. The molecular weights of these bands are 14.9, 15.7 and 17.0 KDa. It was, also observed that total number of bands in snails treated with Basil or Camphor oil was more [15-17 bands] than that of control group [13 bands]

Impact of elevated CO2 concentration under three soil water levels on growth of Cinnamomum camphora / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Forest plays very important roles in global system with about 35% land area producing about 70% of total land net production. It is important to consider both elevated CO(2) concentrations and different soil moisture when the possible effects of elevated CO(2) concentration on trees are assessed. In this study, we grew Cinnamomum camphora seedlings under two CO(2) concentrations (350 micromol/mol and 500 micromol/mol) and three soil moisture levels [80%, 60% and 40% FWC (field water capacity)] to focus on the effects of exposure of trees to elevated CO(2) on underground and aboveground plant growth, and its dependence on soil moisture. The results indicated that high CO(2) concentration has no significant effects on shoot height but significantly impacts shoot weight and ratio of shoot weight to height under three soil moisture levels. The response of root growth to CO(2) enrichment is just reversed, there are obvious effects on root length growth, but no effects on root weight growth and ratio of root weight to length. The CO(2) enrichment decreased 20.42%, 32.78%, 20.59% of weight ratio of root to shoot under 40%, 60% and 80% FWC soil water conditions, respectively. And elevated CO(2) concentration significantly increased the water content in aboveground and underground parts. Then we concluded that high CO(2) concentration favours more tree aboveground biomass growth than underground biomass growth under favorable soil water conditions. And CO(2) enrichment enhanced lateral growth of shoot and vertical growth of root. The responses of plants to elevated CO(2) depend on soil water availability, and plants may benefit more from CO(2) enrichment with sufficient water supply.