| According to the structural features of coumarins with different substituent located atbenzene ring and/or lactone ring, more than40coumarin compounds are classified into4groups and their fluorescent properties are studied. Influences of environmental conditions(pH, solvent polarity, orderly medium, coexisting ions, placing or illumination time, etc.) onfluorescence wavelength and intensity are investigated. Empirical rules of fluorescenceproperty vs. molecular structure are summarized. A fluorimetric method for determination ofHymecromone in Chinese patent medicine Compound Dantong Capsule is proposed, and afluorimetric method as well as a3D fluorescence-chemometrics method for analysis ofNodakenin in Chinese herbal medicine Qianghuo and Kuanyeqianghuo are investigated.Five parts are included in the dissertation.1. Fluorescent properties of coumarins with no substituent on benzene and lactone ringincluding coumarin (benzopyrone) and dihydrocoumarin were investigated. Dilute solution ofcoumarin was no fluorescence under ordinary condition, whereas its alkaline solution canproduce coumarin dimer’s fluorescence with maximum excitation and emission wavelengthsλex/λemat360/495nm and quantum yield (Y)0.09when irradiated by UV light. Addition ofβ-CD into the solution can stabilize the fluorescence. Three dimensional (3D) fluorescencespectra of dihydrocoumarin with λex/λem=272/300nm and Y=0.023were measured, and thespectra was recognized to be similar to substituted benzene. The spectral difference betweencoumarin and dihydrocoumarin illustrate that the presence of substituent on benzene andlactone ring and the cyclic ether oxygen, carbonyl group and ethylenic bond in lactone ringare important to the fluorescence of coumarins.2. Fluorescent properties of8kinds of coumarin with substituent only on lactone ring,including Coumarin-3-carboxylic acid, Ethyl coumarin-3-carboxylate,3-Acetylcoumarin,3-Butyrylcoumarin,4-Hydroxylcoumarin,4-Methoxycoumarin, Warfarin and Dicoumarin,were investigated. The fluorescence of these coumarins is weaker or no fluorescence.Coumarin-3-carboxylic acid produce fluorescence with λex/λem=295/418nm and Y=0.010 only under about pH2.0. Ethylcoumarin-3-carboxylate shows fluorescence with λex/λem=292/418nm and Y=0.009in pH1.9-7.9.4-Hydroxylcoumarin shows fluorescence withλex/λem=285/366nm and Y=0.013in pH5.3-13.5.4-Methoxycoumarin shows very weakfluorescence in pH1.2-12.05. Warfarin shows fluorescence with λex/λem=306/385andY=0.020in pH6.2-13.3.3-Acetylcoumarin,3-Butyrylcoumarin, and Dicoumarin have nofluorescence.3. Fluorescence properties of8kinds of coumarin with substituent only on benzene ring,including6-Methylcoumarin,7-Methylcoumarin, Nodaenin, Xanthotoxin, Psoralen,Isopsoralen,8-Hydroxy-7-methoxycoumarin and8-Acetyl-7-hydroxycoumarin wereinvestigated. The fluorescence properties of these coumarins and more than10kinds of thissort coumarins studied previously in our lab were comprehensively compared, and someempirical rules were obtained as follows:(1) Coumarins with weak electron-donating groupon the benzene ring showed weak fluorescence. For example,6-Methylcoumarin and7-Methylcoumarin have no fluorescence in neutral dilute solution, but give their dimerfluorescence with λex/λem=379/508nm, Y=0.090and λex/λem=355/490nm, Y=0.17,respectively, when pH≥12.20.(2) Unsaturated furan coumarins give weaker fluorescence,while saturated furan and pyran coumarins give stronger fluorescence; fluorescence of linearcoumarins is stronger than that of angle coumarins. For examples, Psoralen is an linerunsaturated furan coumarin, Y=0.029; Isopsoralen is an unsaturated angle furan coumarin, Y=0.015; Praeruptorin is a saturated angle pyran coumarin, Y=0.11; Nodakenin is a saturatedliner furan coumarins, Y=0.81.(3) A-OH or an electron-withdrawing group at ortho-positionof7-OH induce fluorescence decrease, and a-OH at ortho-position of7-OCH3lead tofluorescence quench.(4) Hydrolization of lactone bond and ring-opening under alkalinecondition result in fluorescence quench.4. Fluorescence properties of9kinds of coumarin with substituent both on benzene andlactone ring, including6-Methoxy-4-methylcoumarin,6-Hydroxy-4-methyl coumarin,7-Hydroxy-4-Methylcoumarin,7-Hydroxy-4-(meithoxymethyl) coumarin,7-Acetoxy-4-methylcoumarin,7-methoxy-4-methylcoumarin and4-Methyl-7-ethoxycoumarin, wereinvestigated. Comparison of fluorescent properties of above coumarins leads to some empirical rules as follows:(1) If the substituent of C4position on lactone ring is-CH3,fluorescent property depend on the substituent on benzene ring.(2) If there are multiplesubstituent on both benzene and lactone ring,-OH or-COOH, or stronger electron-donatinggroup or electron-withdrawing group play a decisive role to fluorescence property.(3) Whensubstituent at C6-position is methoxyl and without-OH at any other position, there will be3excitation peaks emerged in fluorescence spectrum.(4) The fluorescence excitationwavelength is associated with the conjugate system of the whole molecule; but thefluorescence emission group is lactone ring.5. A fluorimetric method was proposed for detecting Hymecromone in Chinese patentdrug Compound Dantong Capsule, and the obtained result was consistent with the drug label.A fluorimetric method was also studied for analysis of Nodakenin in Chinese herbal medicineQianghuo and Kuanyeqianghuo, respectively, and HPLC method was used to verify thereliability of the fluorimetric method. The content of Nodakenin in Kuanyeqianghuodetermined by two methods was almost the same,2.94%and3.00%, respectively. However,the results of Nodakenin in Qianghuo were different,0.208%and0.187%, respectively. Thedifference came from the interference of coexistent compounds. For solving the problem, a3D fluorescence-chemometrics method was investigated to determine Nodakenin in Chineseherbal medicine Qianghuo and Kuanyeqianghuo, a satisfied result was obtained, and the resultdemonstrated that3D fluorescence-chemometrics method is an effective method for analysisof complex traditional Chinese medicine. |
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