天然添加物に関わる基礎的調査研究 -その1 天然香料-

2006年厚労省により513種の天然香料其原物質がﾘｽﾄｱｯﾌﾟされた｡当ﾘｽﾄには動物､複合体､生成物､化合物､ｷﾉｺ類､海藻類､樹脂類､規制に関わる物質等が含まれるのでこれらは除いた450種の其原植物について調査研究を遂行することとした｡本ﾌﾟﾛｼﾞｪｸﾄでは天然香料の安全性確認のため以下の調査研究を行った｡1)天然香料に用いられる全ての原料の本体が明確になっていないので､最初に其原植物の学名を決定する｡2)安全性確認のためには食経験の調査が重要なので､食経験に関する文献を広い分野に求めそれらを記述し安全性の評価資料に資する｡3)近年分析機器の急速な発展により多くの成分が単離構造決定されているので､それぞれの其原植物に含まれる成分のﾃﾞｰﾀﾍﾞｰｽを作成する｡4)本ﾃﾞｰﾀﾍﾞｰｽが RTECS ﾃﾞｰﾀﾍﾞｰｽから得られる科学的安全性を支持することを確認する｡The natural resource of flavoring additives 513 items were listed in 2006. Since there are various kinds of complicated materials like animals, complexes, products, components, mushrooms, seaweeds, resins and materials related to the regulations, we removed such items in this project and relisted up 450 items. To make sure the safety of flavoring additives, the following have been investigated. 1) Since the principles of all original plant species using for flavoring additives have not been evident, we confirmed and determined their scientific names individually. 2) The eating experiences as food is important for the confirmation of safety. Therefore, the accumulated scientific data regarding eating experiences published in various fields were incorporated in this project resulting in evaluation of safety. 3) Recently so many natural compounds have been isolated and structurally elucidated depending on the development of purification and structural elucidation technique. We therefore prepared data base of components contained in individual flavoring additive. 4) It becomes evident that the above data base supported the scientific safety data of flavoring additives obtained from RTECS data

ChemInform Abstract: Cephastigiamide A, and Antiplasmodial Activity of Cephalotaxus Alkaloids from Cephalotaxus harringtonia Forma fastigiata.

A new Cephalotaxus alkaloid, cephastigiamide A (1), has been isolated from the leaves of Cephalotaxus harringtonia forma fastigiata and the structure was elucidated by 2D NMR analysis and chemical degradation. Harringtonine, deoxyharringtonine, and homodeoxyharringtonine showed pronounced antiplasmodial activity against Plasmodium falciparum 3D7 but not against Leishmania major. Structure-activity relationship for antiplasmodial activity of isolated Cephalotaxus alkaloids was also discussed

Comparative analysis of transcriptomes in aerial stems and roots of Ephedra sinica based on high-throughput mRNA sequencing

Ephedra plants are taxonomically classified as gymnosperms, and are medicinally important as the botanical origin of crude drugs and as bioresources that contain pharmacologically active chemicals. Here we show a comparative analysis of the transcriptomes of aerial stems and roots of Ephedra sinica based on high-throughput mRNA sequencing by RNA-Seq. De novo assembly of short cDNA sequence reads generated 23,358, 13,373, and 28,579 contigs longer than 200 bases from aerial stems, roots, or both aerial stems and roots, respectively. The presumed functions encoded by these contig sequences were annotated by BLAST (blastx). Subsequently, these contigs were classified based on gene ontology slims, Enzyme Commission numbers, and the InterPro database. Furthermore, comparative gene expression analysis was performed between aerial stems and roots. These transcriptome analyses revealed differences and similarities between the transcriptomes of aerial stems and roots in E. sinica. Deep transcriptome sequencing of Ephedra should open the door to molecular biological studies based on the entire transcriptome, tissue- or organ-specific transcriptomes, or targeted genes of interest

Peptidomic analysis of the venom of the solitary bee Xylocopa appendiculata circumvolans

Abstract Background Among the hymenopteran insect venoms, those from social wasps and bees – such as honeybee, hornets and paper wasps – have been well documented. Their venoms are composed of a number of peptides and proteins and used for defending their nests and themselves from predators. In contrast, the venoms of solitary wasps and bees have not been the object of further research. In case of solitary bees, only major peptide components in a few venoms have been addressed. Therefore, the aim of the present study was to explore the peptide component profile of the venom from the solitary bee Xylocopa appendiculata circumvolans by peptidomic analysis with using LC-MS. Methods A reverse-phase HPLC connected to ESI-OrbiTrap MS was used for LC-MS. On-line mass fingerprinting was made from TIC, and data-dependent tandem mass spectrometry gave MSMS spectra. A major peptide component was isolated by reverse-phase HPLC by conventional way, and its sequence was determined by Edman degradation, which was finally corroborated by solid phase synthesis. Using the synthetic specimen, biological activities (antimicrobial activity, mast cell devaluation, hemolysis, leishmanicidal activity) and pore formation in artificial lipid bilayer were evaluated. Results On-line mass fingerprinting revealed that the crude venom contained 124 components. MS/MS analysis gave 75 full sequences of the peptide components. Most of these are related to the major and novel peptide, xylopin. Its sequence, GFVALLKKLPLILKHLH-NH2, has characteristic features of linear cationic α-helical peptides; rich in hydrophobic and basic amino acids with no disulfide bond, and accordingly, it can be predicted to adopt an amphipathic α-helix secondary structure. In biological evaluation, xylopin exhibited broad-spectrum antimicrobial activity, and moderate mast cell degranulation and leishmanicidal activities, but showed virtually no hemolytic activity. Additionally, the peptide was able to incorporate pores in artificial lipid bilayers of azolectin, confirming the mechanism of the cytolytic activity by pore formation in biological membranes. Conclusions LC-ESI-MS and MS/MS analysis of the crude venom extract from a solitary bee Xylocopa appendiculata circumvolans revealed that the component profile of this venom mostly consisted of small peptides. The major peptide components, xylopin and xylopinin, were purified and characterized in a conventional manner. Their chemical and biological characteristics, belonging to linear cationic α-helical peptides, are similar to the known solitary bee venom peptides, melectin and osmin. Pore formation in artificial lipid bilayers was demonstrated for the first time with a solitary bee peptide