| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1996: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1995: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Research Abstract |
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
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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
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