桂花有效成分合成转化规律与药学研究
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摘要
桂花既是名贵香料植物，也是传统中药材，具有重要研究开发价值。目前，关于桂花资源研究开发的报道很少。本文围绕桂花资源生物学、有效成分合成转化规律及其药学基础开展系统的研究，为高品质桂花资源筛选、采花期的选择、鲜花储藏方式及其加工期的确定，以及桂花资源药用开发及综合利用提供全面的科技支撑。通过系统深入的研究，取得如下创新性研究结果：
(1)首次建立了桂花精油气相色谱指纹图谱，对比不同品种桂花，发现银桂精油含量最高、品质最好，确定银桂为首选优良种质资源。不同品种桂花生态适应性研究表明，丹桂具有较强的生态适应性，是良好的绿化树种。金桂因其氨基酸、微量元素含量相对较高，较适于作为桂花食品资源开发。
(2)首次研究了桂花不同开花期有效成分的合成积累规律，多数桂花赋香成分或药用成分(芳樟醇、γ-癸内酯、β-紫罗兰酮、4-氢-β-紫罗兰酮、棕榈酸、α-亚麻酸)在初花期含量达到最高，确定初花期为桂花最佳采收时期：通过显微分析揭示了桂花香味物质的形成及其分泌过程，花瓣表皮下的薄壁细胞可以形成具有芳香性的精油类物质，并能暂时聚集芳香油类物质，通过表皮细胞的结构以及细胞外壁上的刷状毛散发香味；结合桂花不同开花期生理代谢特征研究，确定了提高桂花有效成分含量的花期养护关键点。
(3)研究了鲜桂花腌制储藏过程中有效成分转化积累规律，确定了桂花储藏40-60天之间进行加工为最佳加工时期。通过研究不同储藏条件对有效成分转化积累的影响，确定了最佳储藏条件为：腌制初期添加0．5％-0．1％柠檬酸，可明显增加桂花浸膏产量和精油品质。
(4)系统研究优化了桂花有效成分分离纯化技术，建立了桂花浸膏、精油及总黄酮的同步制备技术，并实现了中试及产业化应用。其中，桂花浸膏最佳生产工艺为：提取温度24．5℃、提取时间110 min、固液比1：2．7；桂花总黄酮最佳生产工艺为：以1．6倍90％乙醇解析15 min，40倍80％乙醇90℃提取3 h。优选出HPD400大孔树脂纯化桂花总黄酮，建立了相应的纯化工艺：以浓度为0．46 mg／mL样品液上样，吸附流速3 BV／h，上样体积15 BV，洗脱剂为7 BV的70％的乙醇溶液，洗脱流速2 BV／h。纯化后桂花总黄酮纯度和收率分别达90％和80％以上。
(5)首次研究了桂花总黄酮及桂花浸膏的生物活性和药理作用。发现桂花总黄酮具有很好的抑菌活性，对枯草杆菌等的抑菌效果优于目前常用的抑菌剂苯甲酸钠；还发现桂花总黄酮对色拉油等食用油均有较好的抗氧化效果，明显优于BHT等工业抗氧化剂，且与柠檬酸、Vc等协同作用可增强抗氧化效果；通过桂花浸膏和桂花总黄酮对CCl_4引起的急性小鼠肝损伤抑制作用研究表明，桂花浸膏具有极好的抑制肝损伤的效果，肝脏病理切片发现，桂花浸膏大(600 mg／kg)、中(300 mg／kg)、小(150 mg／kg)剂量组对CCl_4引起的肝脏组织的气球样变及细胞性坏死均有很好的治疗效果，优于阳性药物联苯双酯(200 mg／kg)，表明桂花浸膏有望开发为一种保肝、护肝的保健品和药品。
The flower of Osmanthus Fragrans Lour. is of economic importance because it isused as expensive perfumes and traditional medicine. Presently, studies on applicationdevelopment and pharmacy of O. fragrans were rarely reported. In this paper, we studiedthe active components' synthesis conversion and pharmaceutical of O. fragrans. It couldprovide information for people to select the reasonable harvest stages, storage methodsand application of O. fragrans resource. The results of the research were as follows:
(1) GC fingerprint of essential oil of O. fragrans was established using computersoftware. It was effective to evaluate the quality of O. fragrans. Latifolius Group was thebest germplasm resources for perfumery production. We compared contents of aminoacids, trace elements of different varieties of O. fragrans. The result showed thatThunbergii group had better nutritional value and Aurantiaeus group had better ecologicaladaptability.
(2) We studied the rules of synthesis and conversion of essential oil. The resultshowed that the initial flowering stage was the best period for picking. The yield ofessential oil was higher and the main aroma components were also higher at the initialflowering stage. In this paper, we studied the physiological metabolism of O. fragrans andexplained the formation of the fragrance by using optical electron microscopy, scanningelectron microscopy and transmission electron microscopy.
(3) We investigated the changes of the yield and chemical compositions of O.fragrans stored at different conditions. Adding citric acid could significantly increase theyield of essential oil and the best add volume was 0.1%-0.5 %. The result could givesome practical guidance for the utilization of O. fragrans.
(4) The technology of extraction and purification of active components of O.fragrans were studied. Solvent extraction was the best method for essential oil. Theextracting parameters of O. fragrans concrete were determined using Response SurfaceMethododlogy. The best extraction conditions were the temperature of 24.5℃, extractiontime 110 min, the solid-liquid ratio of 1:2.7. The extraction techniques for total flavonoidsof O. fragrans were investigated by contrast. The results showed that release andhot-extraction method was better than other extractions. The optimum purification conditions of total flavonoids with macroporous resin HPD400 were obtained. Withmacroporous resin to adsorb and purify, the yield of total flavonoids was higher than 90 %and the content was higher than 80 %.
(5) The biological activity and pharmacological effects of concrete and totalflavonoids of O. fragrans were studied. We found that total flavonoids of O. fragrans hhigh anti-bacterial and anti-oxidation effects. The lipid antioxidant experiments showedthat total flavonoids of O. fragrans had better anti-oxidation effects in lard, rapeseed oil,soya bean oil, peanut oil. Synergist, such as ascitric acid and Vc, could enhance itsantioxidant effect. With antioxidants, such as BHA, BHT and PG, could enhance itsantioxidant effects.
Effects of concrete and total fiavonoids of O. fragrans on liver injury induced bycarbon tetrachloride in rats were studied. The concrete of O. fragrans had excellent effectof the treatment of liver damage. Liver biopsy revealed that O. fragrans concrete of large(600 mg/kg), medium (300 mg/kg), low-dose (150 mg/kg) groups had excellenttherapeutic effect for balloon cell degeneration and necrosis. It was better than thebiphenyldimethylesterate (200 mg/kg). O. fragrans has the the potential of healthy foodsand drugs.
(1)首次建立了桂花精油气相色谱指纹图谱，对比不同品种桂花，发现银桂精油含量最高、品质最好，确定银桂为首选优良种质资源。不同品种桂花生态适应性研究表明，丹桂具有较强的生态适应性，是良好的绿化树种。金桂因其氨基酸、微量元素含量相对较高，较适于作为桂花食品资源开发。
(2)首次研究了桂花不同开花期有效成分的合成积累规律，多数桂花赋香成分或药用成分(芳樟醇、γ-癸内酯、β-紫罗兰酮、4-氢-β-紫罗兰酮、棕榈酸、α-亚麻酸)在初花期含量达到最高，确定初花期为桂花最佳采收时期：通过显微分析揭示了桂花香味物质的形成及其分泌过程，花瓣表皮下的薄壁细胞可以形成具有芳香性的精油类物质，并能暂时聚集芳香油类物质，通过表皮细胞的结构以及细胞外壁上的刷状毛散发香味；结合桂花不同开花期生理代谢特征研究，确定了提高桂花有效成分含量的花期养护关键点。
(3)研究了鲜桂花腌制储藏过程中有效成分转化积累规律，确定了桂花储藏40-60天之间进行加工为最佳加工时期。通过研究不同储藏条件对有效成分转化积累的影响，确定了最佳储藏条件为：腌制初期添加0．5％-0．1％柠檬酸，可明显增加桂花浸膏产量和精油品质。
(4)系统研究优化了桂花有效成分分离纯化技术，建立了桂花浸膏、精油及总黄酮的同步制备技术，并实现了中试及产业化应用。其中，桂花浸膏最佳生产工艺为：提取温度24．5℃、提取时间110 min、固液比1：2．7；桂花总黄酮最佳生产工艺为：以1．6倍90％乙醇解析15 min，40倍80％乙醇90℃提取3 h。优选出HPD400大孔树脂纯化桂花总黄酮，建立了相应的纯化工艺：以浓度为0．46 mg／mL样品液上样，吸附流速3 BV／h，上样体积15 BV，洗脱剂为7 BV的70％的乙醇溶液，洗脱流速2 BV／h。纯化后桂花总黄酮纯度和收率分别达90％和80％以上。
(5)首次研究了桂花总黄酮及桂花浸膏的生物活性和药理作用。发现桂花总黄酮具有很好的抑菌活性，对枯草杆菌等的抑菌效果优于目前常用的抑菌剂苯甲酸钠；还发现桂花总黄酮对色拉油等食用油均有较好的抗氧化效果，明显优于BHT等工业抗氧化剂，且与柠檬酸、Vc等协同作用可增强抗氧化效果；通过桂花浸膏和桂花总黄酮对CCl_4引起的急性小鼠肝损伤抑制作用研究表明，桂花浸膏具有极好的抑制肝损伤的效果，肝脏病理切片发现，桂花浸膏大(600 mg／kg)、中(300 mg／kg)、小(150 mg／kg)剂量组对CCl_4引起的肝脏组织的气球样变及细胞性坏死均有很好的治疗效果，优于阳性药物联苯双酯(200 mg／kg)，表明桂花浸膏有望开发为一种保肝、护肝的保健品和药品。
The flower of Osmanthus Fragrans Lour. is of economic importance because it isused as expensive perfumes and traditional medicine. Presently, studies on applicationdevelopment and pharmacy of O. fragrans were rarely reported. In this paper, we studiedthe active components' synthesis conversion and pharmaceutical of O. fragrans. It couldprovide information for people to select the reasonable harvest stages, storage methodsand application of O. fragrans resource. The results of the research were as follows:
(1) GC fingerprint of essential oil of O. fragrans was established using computersoftware. It was effective to evaluate the quality of O. fragrans. Latifolius Group was thebest germplasm resources for perfumery production. We compared contents of aminoacids, trace elements of different varieties of O. fragrans. The result showed thatThunbergii group had better nutritional value and Aurantiaeus group had better ecologicaladaptability.
(2) We studied the rules of synthesis and conversion of essential oil. The resultshowed that the initial flowering stage was the best period for picking. The yield ofessential oil was higher and the main aroma components were also higher at the initialflowering stage. In this paper, we studied the physiological metabolism of O. fragrans andexplained the formation of the fragrance by using optical electron microscopy, scanningelectron microscopy and transmission electron microscopy.
(3) We investigated the changes of the yield and chemical compositions of O.fragrans stored at different conditions. Adding citric acid could significantly increase theyield of essential oil and the best add volume was 0.1%-0.5 %. The result could givesome practical guidance for the utilization of O. fragrans.
(4) The technology of extraction and purification of active components of O.fragrans were studied. Solvent extraction was the best method for essential oil. Theextracting parameters of O. fragrans concrete were determined using Response SurfaceMethododlogy. The best extraction conditions were the temperature of 24.5℃, extractiontime 110 min, the solid-liquid ratio of 1:2.7. The extraction techniques for total flavonoidsof O. fragrans were investigated by contrast. The results showed that release andhot-extraction method was better than other extractions. The optimum purification conditions of total flavonoids with macroporous resin HPD400 were obtained. Withmacroporous resin to adsorb and purify, the yield of total flavonoids was higher than 90 %and the content was higher than 80 %.
(5) The biological activity and pharmacological effects of concrete and totalflavonoids of O. fragrans were studied. We found that total flavonoids of O. fragrans hhigh anti-bacterial and anti-oxidation effects. The lipid antioxidant experiments showedthat total flavonoids of O. fragrans had better anti-oxidation effects in lard, rapeseed oil,soya bean oil, peanut oil. Synergist, such as ascitric acid and Vc, could enhance itsantioxidant effect. With antioxidants, such as BHA, BHT and PG, could enhance itsantioxidant effects.
Effects of concrete and total fiavonoids of O. fragrans on liver injury induced bycarbon tetrachloride in rats were studied. The concrete of O. fragrans had excellent effectof the treatment of liver damage. Liver biopsy revealed that O. fragrans concrete of large(600 mg/kg), medium (300 mg/kg), low-dose (150 mg/kg) groups had excellenttherapeutic effect for balloon cell degeneration and necrosis. It was better than thebiphenyldimethylesterate (200 mg/kg). O. fragrans has the the potential of healthy foodsand drugs.
引文
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