【作者】 周倩；

【导师】 纪淑娟；

【作者基本信息】 沈阳农业大学 ， 食品科学与工程， 2014， 博士

【摘要】 蓝莓果实的采收正值高温多雨的季节,由于采后田间热和旺盛的呼吸作用,果实中各种生理代谢加快。冷藏能够延缓果实的后熟衰老过程,但在冷藏后常温货架期中蓝莓果实发生严重的凹陷现象,影响了蓝莓果实的商品品质。因此,研究冷藏蓝莓果蒂凹陷的发生规律及其生理生化机制具有重要意义。本文以“蓝丰”蓝莓为试验材料,通过分析蓝莓果实在采后常温自然后熟过程、冷藏过程以及冷藏后常温货架期中果蒂凹陷的发生情况,探讨蓝莓果蒂凹陷的发生规律。并在此基础上进一步分析凹陷发生与果实活性氧代谢、能量代谢和膜脂代谢的关系,并以其从活性氧代谢、能量代谢和膜脂代谢的角度,深入探讨冷藏后蓝莓果实发生凹陷的生理生化机制,为进一步探索调控措施提供理论依据。其主要研究结果如下：冷藏能够有效延长蓝莓果实的贮藏保鲜期,但经长期冷藏的蓝莓果实转入常温货架期后,果实品质下降,在货架期中表现出明显的凹陷症状。而这种凹陷症状在蓝莓果实常温贮藏过程中和冷藏过程中均无明显症状。通过对蓝莓果实冷藏过程以及冷藏后货架期中形态特征以及超微结构的分析表明,随着蓝莓果实冷藏后货架期时间的延长,蓝莓果蒂凹陷的程度加重。通过果实的剖面图分析显示,凹陷蓝莓果实出汁率增加,果肉伴有红变现象发生。细胞的超微结构显示,果实中细胞壁的外层结构被破坏,纤维丝逐渐减少。细胞膜、细胞核膜以及原生质膜发生了不同程度的损伤,淀粉粒减少甚至丧失,这时在蓝莓果实的外观上表现为根蒂部凹陷的发生。从细胞水平上阐述了蓝莓果实冷藏后凹陷发生对细胞超微结构的影响。蓝莓果蒂凹陷发生时伴随着一系列品质的变化,其中包括蓝莓果实硬度、Vc含量的明显下降、腐烂率的上升,糖酸比失调等,果实失去了原有的风味与品质。其冷藏时间越长,货架期所表现的凹陷症状越明显,果实品质下降越严重。通过对苯丙氨酸解氨酶(PAL)和过氧化物酶(POD)两种抗逆境相关酶的分析可以看出,随着果蒂凹陷现象的发生,其PAL出现异常的上升而POD异常下降。通过对各指标在不同贮藏过程中的分析可以看出,蓝莓果实的凹陷有可能是因为长时间的低温逆境胁迫导致。冷藏过程中,蓝莓果实·O2-产生速率、H2O2含量的增加以及SOD、CAT和APX活性的下降均受到了一定程度的抑制。但当果实由冷藏转入常温货架期后,果实的活性氧代谢呈现了一定的差异。随着冷藏后货架期中果蒂凹陷的发生,蓝莓果实·O2-产生速率、H2O2含量急剧增加,SOD、CAT和APX活性迅速下降,出现了异常的变化趋势。通过进一步的相关性分析表明蓝莓果实在冷藏后货架期中·O2-产生速率、H2O2含量的增加以及SOD、CAT、APX活性下降及异常变化是导致蓝莓果蒂凹陷发生的重要原因之一。冷藏蓝莓果实发生凹陷时,能量物质ATP、ADP含量及能荷水平下降迅速,膜脂的过氧化作用不断加剧。冷藏时间越长凹陷发生程度越严重,蓝莓果实ATP、ADP含量和能荷值越低,细胞能量亏缺越严重,线粒体呼吸代谢相关酶H+-ATPase、Ca2+-ATPase以及SDH和CCO活性下降迅速。这些酶活性的异常降低导致了细胞能量的大量亏缺,抑制了线粒体能量的不断合成,进而使细胞膜完整性遭到破坏,酶与底物开始接触。这些结果说明,较高的能量水平可以有效保护细胞膜的完整性,蓝莓果实冷藏后货架期中凹陷发生与能量水平密切相关。通过对蓝莓果实中脂肪酸的种类和相对含量分析发现,蓝莓果实中脂肪酸的种类和含量丰富,不饱和脂肪酸含量高达67.598%。蓝莓果实在冷藏过程中,脂肪酸的组成未发生明显的变化,而不饱和脂肪酸的相对含量、脂肪酸的不饱和指数和不饱和度呈逐渐下降趋势。蓝莓果蒂凹陷发生后,细胞膜透性急剧增大,MDA含量升高显著,膜脂过氧化现象严重。通过相关性分析表明,冷藏蓝莓果蒂凹陷发生与细胞膜脂组分含量、不饱和度以及膜脂过氧化密切相关。蓝莓果实在冷藏后货架期中不饱和脂肪酸含量的下降以及细胞膜透性、脯氨酸含量增加以及LOX活性下降及异常变化是导致蓝莓果蒂凹陷发生的又一重要原因。低温预贮处理可以有效减轻冷藏后货架期中蓝莓果蒂凹陷的发生。使其保持了较高的果实硬度和Vc含量；抑制了果实组织细胞膜透性的增加以及膜脂过氧化产物的积累；提升了蓝莓果实贮藏过程中活性氧代谢相关酶(SOD、CAT和APX)的活性,维持了果实较高的自由基及活性氧清除能力。同时低温预贮处理通过调节线粒体呼吸代谢酶的活性,始终使果实维持一个相对较高的能量水平,防止了采后果实因能量亏损而导致的生理伤害。从而进一步证实了蓝莓果蒂凹陷与活性氧代谢、膜脂代谢和能量代谢密切相关。更多还原

【Abstract】 Blueberry fruits are harvested at high temperature and rainy season. Physiological metabolism of fruits speeds up after picked up because of the filed heat and the hearty respiration. Ripening and senescence have been delayed during the cold storage. But pitting developed severely under improper refrigeration, especially during shelf life after cold storage. The quality of commodity of blueberry fruits has been affected. Therefore, it’s significantly that studied the regularity and physiology and biochemistry mechamism accompanying pitting development. Blueberries(Vaccinium spp.’LanFeng’) are the experimental material in this paper. The objective of this research was to discuss the regulatity of pitting in blueberries by analyzing the occurrence of piting during postharvest natural-rippening, cold storage and shelf-life after cold storage. And the relationship between pitting and reactive oxygen metabolism, energy metabolism and membrane lipid metabolism was explored. On this basis, the physiology and biochemistry mechamism accompanying pitting development has been in-depth exploration, which provides the theoretical foundation for the control measures of storage of blueberries. The main research results are as follows:The storage life of blueberries was extended at low temperature, but fruits quality declined during shelf-life after long-term cold storage. And the pitting symptom showed up obviously at the shelf-life. The fruits stored directly at room temperature and low temperature showed no problem. The analysis, which is the morphological characteristics and ultrastructure during the cold storage and the shelf-life after cold storage, shows that the pitting of blueberries was aggravated as the shelf-life time went on. Profile map of fruits displays that juice yield increased and pulpa pinked accompanying pitting development. At the same time, the cellosilk was reduced and the outer structure was damaged in cell wall. Different levels of injure has been happened in cytomembrane, caryotheca and plasmalemma. The starch grain reduced even losed. The apparent changes show that pitting development in the root of fruits. The effect of pitting development accompanying cell ultrastructure has been stated at cellular level.The qualities of blueberries changed accompanying pitting development including low firmness and Vc content, rising decay incidence and the imbalance sugar-acid ratio, which made the original relish and quality losed. The symptom of pitting expressed more obviouse and the quality declined more serious along with the extended time of cold storage. Phenylalanine ammonia lyase (PAL) and peroxidase (POD) are key enzymes involved in plant stress responses. By analyzing the activity of PAL and POD, it’s concluded that the activity of PAL increased and POD declined abnormally accompanying pitting development of blueberries. The long-term cold stress may be the reason of pitting development of blueberries according to analyzing each index during different storage process. During the cold storage, the increased of·O2-producing rate and H2O2content, and the decreased of activity of SOD, CAT and APX were inhibited at a certain degree. However, the reactive oxygen metabolism appeared the difference when blueberries were transferred from low temperature to the shelf-life at room temperature. Blueberries stored at shelf-life after cold storage when pitting occurred had higher-02producing rate and H2O2content, and lower activity of antioxidant-related enzymes, including superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APX), than those stored at room temperature. It appeared unusual variation trend. The correlation analysis showed that one of the important reason of pitting development in blueberries at shelf-life after cold storage was the increased of·O2producing rate and H2O2content, the decreased of activity of SOD, CAT and APX and the anomalous changes.Contents of ATP, ADP and EC level rapidly decreased as pitting development in blueberries while the membrane peroxidation aggravated. The occurance degree of pitting generally aggravated as storage time progressed, at the same time, contents of ATP, ADP and EC were lower, energy loss of the cell were serious, and the activities of mitochondria respiratory metabolism-related enzymes-H+-ATPase, Ca2+-ATPase, SDH and CCO decreased rapidly. These changes could lead to energy loss of the cell and inhibited the synthesis of energy. The cell membrane integrity was destroyed. Enzyme and substrate began to contact. These results presented here demonstrated the high energy level could protect the integrity of cell membrane and alleviate the pitting development and the closely relationship between pitting and energy level by exposing blueberries to shelf-life after cold storage.The blueberries are rich in kind and content of fatty acid by analyzing the kind and content of fatty acid. The relative percentage content of unsaturated fatty acid was67.598%. During the cold storage, the composition of fatty acid has not changed obviously, but the relative content of unsaturated fatty acid, unsaturation index and degree of unsaturation of fatty acid decreased gradually as the extending of the storage time. The cell membrane permeability increased sharply, the MDA content rised significantly and the membrane lipid peroxidation was severe accompanying pitting development. The correlation analysis showed that the closely relationship between pitting and content of membrane lipid composition, degree of unsaturation and the peroxide of cytomembrane. Another important reasons of pitting development in blueberries at shelf-life after cold storage was the decreased of unsaturated fatty acid content and the LOX activity, the increased of cell membrane permeability and proline content.LTC treatment effectively alleviated the pitting development in blueberries, maintained the higher firmness and Vc content, and inhibited the increase of cell membrane permeability and the accumulation of product of lipid peroxidation during shelf-life after cold storage. Meanwhile, it promoted the activity of reactive oxygen metabolism related enzyme (SOD, CAT and APX), maintained higher scavenging ability on free radical and reactive oxygen. The treatment always maintains the higher energy levels and prevents the physiology damage of blueberries due to the energy loss by regulating the activities of mitochondria respiratory metabolism-related enzymes. These results were furtherly confirmed the closely relationship between pitting and reactive oxygen metabolism, membrane lipid metabolism and energy metabolism.更多还原