【作者】 于洋；

【导师】 杨国慧；

【作者基本信息】 东北农业大学 ， 果树学， 2014， 硕士

【摘要】 树莓为蔷薇科(Rosaceae)悬钩子属(Rubus.L)小浆果果树,具有极高的食用药用价值。本研究以4个果实颜色不同的树莓品种(紫色果：美22、红色果：欧洲红、黄色果：金秋、黑树莓)为试材,分别于4个时期：5月末、6月末、7月末、8月末采集结果枝和根蘖苗上的叶片,以及4个品种的青果和成熟的果实。采用有机溶剂结合超声波辅助法,通过正交试验设计确定了叶片和果实中的黄酮类化合物的提取方法,并采用NaNO2-Al(NO3)3比色法测定其含量。分析了树莓黄酮类化合物的稳定性以及对4种自由基(羟自由基、超氧阴离子自由基、ABTS自由基、DPPH自由基)的清除能力,利用打孔法和菌落记数法测定了其对4种真菌和3种细菌的抑制能力。通过分析不同种类、不同发育时期树莓叶片和果实中黄酮类化合物的含量及其生理活性,为筛选高含量的黄酮类化合物树莓种质资源和生产树莓保健产品提供科学依据。通过试验得出结论如下：1.利用超声波辅助法从树莓叶片中提取黄酮类化合物的最佳提取条件是：95%的乙醇溶液在100W的超声辅助条件下提取1.5h,料液比为1:12(g/mL);从树莓果实中提取黄酮类化合物的最佳提取条件是：95%的乙醇溶液在100W的超声辅助条件下提取1.5h,料液比为1：10(g/mL)。2.4个树莓品种叶片黄酮类化合物均在6月末表现最高,但在相同时期4个品种含量高低次序不同,在6月末为欧洲红>美22>金秋>黑树莓,在同一个发育时期,欧洲红和美22的结果枝叶片要高于根蘖苗叶片；4个品种青果中的黄酮类化合物含量均显著高于熟果中的含量,且不论熟果还是青果,4个品种的高低次序均为金秋>美22>欧洲红>黑树莓；同一品种树莓,其叶片黄酮含量高于果实。3.树莓叶片黄酮类化合物对4种自由基清除力与其含量呈正相关,6月末清除力最强,4个品种的清除能力为欧洲红>美22>金秋>黑树莓,同一品种对不同自由基清除力为DPPH>ABTS>羟自由基>超氧阴离子；树莓果实清除力亦与其黄酮含量呈正相关,青果清除力均大于熟果,青果对4种自由基清除力相当,而熟果对ABTS、DPPH的清除力要高于羟自由基和超氧阴离子。4.树莓叶片黄酮类化合物对纹枯病菌的抑制能力和其含量呈正相关,6月末抑制能力最强,美22树莓叶片提取物对4种真菌的抑制能力表现为黑斑病菌>纹枯病菌>根腐病菌>枯萎病菌；不同成熟度果实对对纹枯病菌的抑制能力和其含量表现为负相关,成熟果实均高于青果,品种金秋熟果提取物对4种真菌的抑制能力为黑斑病菌>纹枯病菌>枯萎病菌>根腐病菌,青果提取物对4种真菌的抑制能力为黑斑病菌>枯萎病菌>根腐病菌>纹枯病菌。5.树莓叶片黄酮类化合物对大肠杆菌抑制能力和其含量表现为正相关,6月末抑制能力最强,美22树莓叶片提取物对3种细菌的抑制能力表现为枯草芽孢杆菌>金黄色葡萄球菌>大肠杆菌；不同成熟度的果实对大肠杆菌的抑制能力和其含量表现为负相关,熟果均高于青果,金秋果实提取物对3种细菌的抑制能力表现为枯草芽孢杆菌>金黄色葡萄球菌>大肠杆菌。6.树莓叶片黄酮类化合物在80℃范围内表现稳定；具有一定程度的耐酸性,其在pH5.0-8.0之间表现稳定；在常温下贮存,随着贮存时间的加长,树莓叶黄酮表现出较快的降解,在避光条件下保存36h提取液吸光度降低的幅度也达到了23.3%;还原剂Na2SO3及柠檬酸对树莓叶黄酮稳定性均有一定程度的降低作用,而蔗糖和NaCl在浓度小于0.25%时对其无影响。更多还原

【Abstract】 Raspberry, a small berry fruit tree, which belongs to Rosaceae Rubus. Land has high medical and edible value. Four different colors of raspberries (purple fruit:America-22, red fruit:Europe-red, yellow fruit:Fall gold, black raspberries) were used as test materials in this study. The leaves of fertile branch and root sucker seedling were collected in four period, respectively at the end of may, June, July, and August, and the green and ripe fruits of four varieties were also collected. The organic solvent in combination with ultrasonic auxiliary method was used, through the contents of orthogonal experiment design, the method of extracting the flavonoids and the NaNO2-Al(N03)3colorimetric method were used to measure the contents of flavonoids from leaves and fruits. The stability and the remove power for four kinds of free radicals (OH·, O2·, ABTS·, DPPH·) on raspberry flavonoids were analyzed, and the inhibition capacity for four kinds of fungi and three kinds of bacteria were measured using the method of punching and colony notation. The analysis of flavonoids content and physiological activity of different types and different development period of raspberry leaf and fruit would provide a scientific basis for screening high levels of flavonoids in raspberry germplasm resources and raspberry health care products. Through conclusion of the test is obtained as follows:1. By using ultrasonic assisted method, the best extraction conditions of extracting flavonoids from raspberry leaf and fruit was:95% ethanol,100W ultrasonic assisted extracting for1.5h, the ratio of material:liquid was1:12(g/mL).2. The contents of flavonoids in raspberry leaf of four varieties all reached the maximum in late June. But in different periods, the contents of of flavonoids in four varieties showed different order. In late June, Europe-red> America22> Fall gold> black raspberries. In the same growth period, in Europe-red and America22, the contents of flavonoids in leaf of fertile branch leaf is higher than root sucker seedling. The contents of flavonoids in green fruit were significantly higher than in ripe fruit in four varieties. Both green fruit and ripe fruit, the contents of flavonoids in four varieties had the same order:Fall gold> America22> Europe-red> black raspberries. In the same kind raspberry, the content of flavonoids in leaf is higher than in fruit.3.The remove power of flavonoids in raspberry leaf for OH·, ABTS·, DPPH·, O2·showed positive correlation with its content, the content was more higher, the remove power was more stronger and which reached the peak in late June. The the remove power of different radical scavenging is DPPH·> ABTS·> OH·> O2-·.The remove power of flavonoids in raspberry fruit also showed positive correlation with its content, and the power in green fruit is greater than in ripe fruit. Among them, the remove power for OH·and O2-· of green fruit was much higher than ripe fruit. The remove power of green fruit for four kinds of free radicals is almost equal, however, the remove power of ripe fruit for ABTS·and DPPH·were higher than OH·and O2-·. For the same raspberry, the remove power of extraction for free radicals of fruit is higher than leaf.4. The inhibition ability of flavonoids in raspberry leaf for Rhizoctorzia solani showed positive correlation with its content, showing June> July> August> May as a whole. The inhibition ability of leaf extracting in June for four kinds of fungi showed:Alternaria brassicicola> Rhizoctorzia solani> Root rot fungus> oxysporum. The inhibition ability of flavonoids in raspberry fruit for Rhizoctorzia solani showed negative correlation with its content and the inhibition ability on four varieties ripe fruit is greater than green fruit. Fall gold ripe fruit extracting performance for Alternaria brassicicola> Rhizoctorzia solani> oxysporum> Root rot fungus, however, the inhibition ability for four kinds of fungi of Fall gold green fruit extracting performance for Alternaria brassicicola>Root rot fungus>oxysporum>Rhizoctorzia solani.5. The inhibition ability of flavonoids in raspberry leaf for escherichia coli showed positive correlation with its content.showing June> July> August> May. The inhibition ability of America-22leaf extracting in June for three kinds of bacteria showed that Bacillus subtilis> Staphylococcus aureus> escherichia coli. The inhibition ability of flavonoids in raspberry leaf for escherichia coli showed negative correlation with its content,and the inhibition ability of four varieties ripe fruit is greater than green fruit.Fall gold fruit extracting for three kinds of bacteria performance for Bacillus subtilis> Staphylococcus aureus> escherichia coli.6. The flavonoids of raspberry leaf was stable at scope of80℃, which had a certain degree of resistance to acid and a stable performance between pH5.0-8.0. In room temperature, as the continuity of storage time, the raspberry leaf flavonoids showed faster degradation. Under the condition of dark for36h, the extracting absorbance decrease also reached23.3%. Reductant Na2SO3and citric acid has a certain degree influence on raspberry leaf flavonoids stability, but the concentration of sucrose and NaCl in less than0.25%had no effect on it.更多还原