Bioorganic Studies on the Fragrance Formation during Flower Opening.

在花期香味形成的研究过程中，我们提纯了栀子和茉莉的酒精香气前体。这些糖苷的分离采用粗酶测定法。从这些花蕾中分离出的所有醇类香气前体均为β-香草苷、β-月见草苷、β-芦丁苷和6‘-O-丙二酰基-β-D-吡喃葡萄糖苷。除了这些醇香，还从玫瑰花中分离出了几种香叶基和香茅基二糖苷。这是首次从月季花中分离到单萜醇的二糖苷类化合物。为了阐明醇类香气的形成机理，我们试图从开花期采摘的茉莉花中制备和纯化催化醇类香气苷水解酶。通过监测酶对Rho-硝基苯基-β-D-谷氨酸…的水解活性来确定酶的活性更多的吡喃葡萄糖苷(Ro NP-GLC)和Ro NP-Primeveroside(Ro NP-PRI)。从开花阶段获得的丙酮粉在柠檬酸缓冲液(pH 6.0)中增溶。上清液用冷丙酮沉淀，然后用硫酸铵沉淀。得到的蛋白质在CM-Toyota opearl 650M色谱柱上层析，得到葡萄糖苷酶I和葡萄糖苷酶II，以及Primeverosidase I。Primeverosidase I进一步经CM-Toyota opearl柱上的FPLC层析得到单峰。用该酶对RhoNP-PRI进行酶解，并对其水解物进行薄层分析，未检测到木糖和葡萄糖。在水解液中只检测到青蒿素是唯一的糖类。这一结果强烈地表明，该酶像内切-β-葡萄糖苷酶一样，一步地将酒精性香气中的二糖糖苷水解，释放出酒精性香气和二糖。为了检测凝胶上的低水平糖苷酶活性，首次合成了具有荧光的底物--4-甲基伯菲尔基二糖糖苷。利用香气传感器和微波炉提取，可以监测开花过程中香气的释放和酒精类香气前体含量的变化。较少

In the course of the studies on the fragrance formation during flower opening, we have purified alcoholic aroma precursors of Gardenia jasminoides and Jasminum sambac. The isolation of these glycosides was guided by the crude enzyme assay. All the alcoholic aroma precursors isolated from these flower buds were beta-vicianoside, beta-primeveroside, beta-rutinoside, and 6'-O-malonyl-beta-D-glucopyranoside. Besides these alcoholic aromas, several geranyl and citronellyl disaccharide glycosides have been isolated-from rose flowers. This is for the first time that the disaccharide glycosides of monoterpene alcohols have been isolated from rose flowers.In order to clarify the mechanism of alcoholic aroma formation, we tried to prepare and purify the enzymes catalyzing the hydrolysis of the alcoholic aroma glycosides from the flowers of G.jasminoides flowers picked at the opening stage. The enzyme activity was determined by monitoring the hydrolysis activity toward rho-nitrophenyl beta-D-gluc … More opyranoside (rhoNP-Glc) and rhoNP-primeveroside (rhoNP-Pri). The acetone powder obtained from the flowers at the opening stage was solubilized in citrate buffer (pH 6.0). The supernatant was subjected to the precipitation with cold acetone and then by ammonium sulfate. The protein thus obtained was chromatographed on a CM-Toyopearl 650M column to give Glucosidases I and II,and Primeverosidase I.The Primeverosidase I was further subjected to FPLC on CM-Toyopearl column to give a primeverosidase as a single peak. rhoNP-Pri was hydrolyzed by the primeverosidase and the resultant hydrolysate was analyzed by TLC.Neither xylose nor glucose was detected. Only primeverose was detected as the sole saccharide in the hydrolysate. This result strongly suggested that the primeverosidase hydrolyzed disaccharide glycoside of slcoholic aromas like an endo-beta-glucosidase in a single step to liberate alcoholic aroma and disaccharide. To detect the low level of glycosidase activities on the gel after electro-phoreisis, the substrates with fluorecsence, 4-methylumberifellyl disaccharide glycosides, were synthesized for the first time.The use of aroma sensor and microwave oven-extraction has made it possible to monitor the aroma release and the change in the contents of alcoholic aroma precusors during flower opening.The results obtained so far have given important information and prompt further studies on the subject. Less