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.
Cinnamomum camphora/enzymology* ; Alkyl and Aryl Transferases/chemistry*

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.
Anti-Inflammatory Agents/pharmacology* ; Cinnamomum ; Cinnamomum camphora ; Plant Leaves

OBJECTIVETo measure and compare the content of d-borneol in the different parts of Cinnamomum camphora by GC-MS.

METHODWith water-steam distillation and GC-MS method, d-borneol was extracted and determined.

RESULTThe 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%.

CONCLUSIONThe 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.

Bornanes ; analysis ; Cinnamomum camphora ; chemistry ; Gas Chromatography-Mass Spectrometry ; methods ; Plant Extracts ; analysis ; Plant Structures ; chemistry

OBJECTIVETo optimize the condition of callus of Cinnamonum camphora induced.

METHODGC and plant tissue culture method were applied in the study.

RESULTThe 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.

CONCLUSIONThe 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.

Bornanes ; metabolism ; Cinnamomum camphora ; growth & development ; metabolism ; Culture Media ; Tissue Culture Techniques

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.
Cinnamomum camphora/genetics* ; Cloning, Molecular ; Genes, Plant ; Phosphotransferases (Phosphate Group Acceptor)/genetics* ; Phylogeny ; Sequence Alignment

OBJECTIVETo identify endophytic fungi bn12 from Cinnamomum camphora chvar, borneol and analysis its volatile metabolites.

METHODThe endophytic fungi bn12 was identified by morphological observation. volatile metabolites of endophytic fungi bn12 was analyzed by gas chromatography/mass spectrography (GC-MS).

RESULTVolatile 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.

CONCLUSIONEndophytic fungi bn12 is belong to pleosporaceae fungus. It has the ability of producing broneol.

Bornanes ; analysis ; Cinnamomum camphora ; microbiology ; DNA, Ribosomal Spacer ; genetics ; Endophytes ; genetics ; metabolism ; Fungi ; classification ; genetics ; metabolism ; Gas Chromatography-Mass Spectrometry ; Phylogeny ; Volatile Organic Compounds ; analysis

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.
Biomass ; Carbon Dioxide ; metabolism ; Cinnamomum camphora ; growth & development ; metabolism ; Plant Roots ; growth & development ; metabolism ; Plant Shoots ; growth & development ; metabolism ; Soil ; Trees ; growth & development ; metabolism ; Water ; metabolism

OBJECTIVETo 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.

METHODThe 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.

RESULTAs 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.

CONCLUSIONThe 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.

Alkanes ; therapeutic use ; Animals ; Anti-Inflammatory Agents ; therapeutic use ; Arthritis ; chemically induced ; drug therapy ; metabolism ; Body Weight ; drug effects ; Bornanes ; therapeutic use ; Cinnamomum camphora ; chemistry ; Freund's Adjuvant ; adverse effects ; Interleukin-2 ; metabolism ; Interleukin-6 ; metabolism ; Male ; Random Allocation ; Rats ; Rats, Wistar ; Tumor Necrosis Factor-alpha ; metabolism