线粒体分离纯化及其在果实成熟衰老研究中的应用进展

doi:10.16420/j.issn.0513-353x.2023-0528

摘要/Abstract

摘要：

综述了果实线粒体分离纯化的方法、线粒体制备的难点和关键点、线粒体纯度和完整性的评价方法以及制备线粒体在果实成熟衰老研究中的应用，并探讨了存在的问题及未来的发展方向。

Abstract:

This paper reviews the recent advances in mitochondrial isolation and purification and relevant application in research on fruit ripening and senescence，as well as the difficulties and crucial factors in mitochondrial preparation, purity and intactness evaluation of isolated mitochondria. The existing problems and developing trend in the future are also discussed.

Key words: fruit, mitochondria, isolation and purification, ripening and senescence

李欣, 柴应芳, 田真, 王鹏蔚, 程运江. 线粒体分离纯化及其在果实成熟衰老研究中的应用进展[J]. 园艺学报, 2024, 51(7): 1516-1528.

LI Xin, CHAI Yingfang, TIAN Zhen, WANG Pengwei, CHENG Yunjiang. Isolation and Purification of Mitochondria and Its Application in Research on Fruit Ripening and Senescence[J]. Acta Horticulturae Sinica, 2024, 51(7): 1516-1528.

图/表 3

参考文献 86

[1]	Ali M, Raza M A, Li S, Huan C, Zheng X. 2020. 1-Methylcyclopropene treatment controls ethanol accumulation associated with regulation of mitochondrial energy metabolism in kiwifruit(Actinidia deliciosa)cv.“Bruno”during storage at room temperature. Journal of Food Biochemistry, 44 (7):e13273.
[2]	Almeida A M, Navet R, Jarmuszkiewicz W, Vercesi A E, Sluse-Goffart C M, Sluse F E. 2002. The energy-conserving and energy-dissipating processes in mitochondria isolated from wild type and nonripening tomato fruits during development on the plant. Journal of Bioenergetics and Biomembranes, 34 (6):487-498. doi: 10.1023/a:1022574327117 pmid: 12678440
[3]	Baqui S M, Mattoo A K, Modi V V. 1974. Mitochondrial enzymes in mango fruit during ripening. Phytochemistry, 13 (10):2049-2055.
[4]	Biale J B, Young R E, Popper C S, Appleman W E. 1957. Metabolic processes in cytoplasmic particles of the avocado fruit. I. Preparative procedure, cofactor requirements,and oxidative phosphorylation. Physiologia Plantarum, 10 (1):48-63.
[5]	Bogin E, Erickson L C. 1965. Activity of mitochondrial preparations obtained from Faris sweet lemon fruit. Plant Physiology, 40 (3):566-569. doi: 10.1104/pp.40.3.566 pmid: 16656127
[6]	Boussardon C, Przybyla-Toscano J, Carrie C, Keech O. 2020. Tissue-specific isolation of Arabidopsis/plant mitochondria - IMTACT(isolation of mitochondria tagged in specific cell types). The Plant Journal, 103 (1):459-473. doi: 10.1111/tpj.14723 pmid: 32057155
[7]	Cai J, Wang P, Tian S, Qin G. 2018. Quantitative proteomic analysis reveals the involvement of mitochondrial proteins in tomato fruit ripening. Postharvest Biology and Technology, 145:213-221.
[8]	Chotikakham S, Panya A, Saengnil K. 2022. Methyl salicylate treatment alleviates peel spotting of‘Sucrier’banana by improving mitochondrial physiological properties and functions. Postharvest Biology and Technology, 186:111832.
[9]	Costa A D T, Nantes I L, Ježek P, Leite A, Arruda P, Vercesi A E. 1999. Plant uncoupling mitochondrial protein activity in mitochondria isolated from tomatoes at different stages of ripening. Journal of Bioenergetics and Biomembranes, 31 (5):527-533. doi: 10.1023/a:1005408809619 pmid: 10653480
[10]	de Oliveira J G, Morales L M M, Silva G M C, da Cruz Saraiva K D, Santana D B, dos Santos C P, Oliveira M G, Costa J H. 2017. Procedures of mitochondria purification and gene expression to study alternative respiratory and uncoupling pathways in fruits. //Jagadis Gupta K. Plant Respiration and Internal Oxygen. New York: Humana Press.
[11]	de Virville J D, Diolez P, Moreau F, Dessaux C, Marcellin P. 1987. Oxidative and phosphorylating activities of mitochondria isolated from tomato fruit during the post-chilling burst of respiration. Journal of Plant Physiology, 129 (3-4):341-349.
[12]	Dickinson D B, Hanson J B. 1965. Comparison of mitochondria from tomato fruits at various stages of ripeness. Plant Physiology, 40 (1):161-165. doi: 10.1104/pp.40.1.161 pmid: 16656058
[13]	Dickinson D B, Misch M J, Drury R E. 1970. Freezing damage to isolated tomato fruit mitochondria as modified by cryoprotective agents and storage temperature. Plant Physiology, 46 (2):200-203. doi: 10.1104/pp.46.2.200 pmid: 16657434
[14]	Haard N F, Hultin H O. 1968. An improved technique for the isolation of mitochondria from plant tissue. Analytical Biochemistry, 24 (2):299-304. pmid: 5671028
[15]	Haard N F, Hultin H O. 1969. Abnormalities in ripening and mitochondrial succinoxidase resulting from storage of preclimacteric banana fruit at low relative humidity. Phytochemistry, 8 (11):2149-2152.
[16]	He Fang, He Dacheng. 2005. The development of the techniques of subcellular isolation. Modern Instruments,(2):1-5. (in Chinese)
	贺芳, 何大澄. 2005. 亚细胞器分离纯化技术的发展. 现代仪器,(2):1-5.
[17]	Hobson G E. 1969. Respiratory oxidation by mitochondria from tomatoes at different stages of ripeness. Qualitas Plantarum et Materiae Vegetabiles, 19:155-165.
[18]	Hobson G E, Lance C, Young R E, Biale J B. 1966. Isolation of active subcellular particles from avocado fruit at various stages of ripeness. Nature, 209:1242-1243.
[19]	Holtzapffel R C, Finnegan P M, Millar A H, Badger M R, Day D A. 2002. Mitochondrial protein expression in tomato fruit during on-vine ripening and cold storage. Functional Plant Biology, 29 (7):827-834. doi: 10.1071/PP01245 pmid: 32689530
[20]	Hulme A C, Jones J D, Wooltorton L S C. 1964. Mitochondrial preparations from the fruit of the apple-I. Phytochemistry, 3 (2):173-188.
[21]	Jarmuszkiewicz W, Almeida A M, Sluse-Goffart C M, Sluse F E, Vercesi A E. 1998. Linoleic acid-induced activity of plant uncoupling mitochondrial protein in purified tomato fruit mitochondria during resting,phosphorylating, and progressively uncoupled respiration. The Journal of Biological Chemistry, 273 (52):34882-34886.
[22]	Jeffery D, Goodenough P W, Weitzman P D J. 1986. Enzyme activities in mitochondria isolated from ripening tomato fruit. Planta, 168:390-394. doi: 10.1007/BF00392366 pmid: 24232150
[23]	Jin P, Zhu H, Wang J, Chen J, Wang X, Zheng Y. 2013. Effect of methyl jasmonate on energy metabolism in peach fruit during chilling stress. Journal of the Science of Food and Agriculture, 93 (8):1827-1832. doi: 10.1002/jsfa.5973 pmid: 23208649
[24]	Jin P, Zhu H, Wang L, Shan T, Zheng Y. 2014. Oxalic acid alleviates chilling injury in peach fruit by regulating energy metabolism and fatty acid contents. Food Chemistry, 161:87-93. doi: 10.1016/j.foodchem.2014.03.103 pmid: 24837925
[25]	Jin Tian, Xu Yuemei, Kuang Guanling, Liu Guidong. 2024. Effect of boron deficiency on the root growth and mitochondrial function of trifoliate orange seedlings. Acta Horticulturae Sinica, 51 (1):121-132. (in Chinese) doi: 10.16420/j.issn.0513-353x.2022-1116
	金天, 徐月美, 邝冠翎, 刘桂东. 2024. 缺硼胁迫对枳幼苗根系生长及线粒体功能的影响. 园艺学报, 51 (1):121-132. doi: 10.16420/j.issn.0513-353x.2022-1116
[26]	Jing G, Zhou J, Zhu S. 2016. Effects of nitric oxide on mitochondrial oxidative defence in postharvest peach fruits. Journal of the Science of Food and Agriculture, 96 (6):1997-2003. doi: 10.1002/jsfa.7310 pmid: 26084831
[27]	Jones J D, Hulme A C. 1961. Preparation of mitochondria from the peel of apples. Nature, 191:370-372.
[28]	Kalra S K, Brooks J L. 1973. Lipids of ripening tomato fruit and its mitochondrial fraction. Phytochemistry, 12 (3):487-492.
[29]	Kalra S K, Brooks J L. 1979. Behaviour of some mitochondrial enzymes in ripening tomato fruit. Phytochemistry, 18 (2):2017-2019.
[30]	Kan J, Wang H, Jin C. 2011. Changes of reactive oxygen species and related enzymes in mitochondrial respiration during storage of harvested peach fruits. Agricultural Sciences in China, 10 (1):149-158.
[31]	Kan J, Wang H, Jin C, Xie H. 2010. Changes of reactive oxygen species and related enzymes in mitochondria respiratory metabolism during the ripening of peach fruit. Agricultural Sciences in China, 9 (1):138-146.
[32]	Kane O, Marcellin P. 1978. Incidence of ripening and chilling injury on the oxidative activities and fatty acid compositions of the mitochondria from mango fruits. Plant Physiology, 61 (4):634-638. doi: 10.1104/pp.61.4.634 pmid: 16660352
[33]	Katz E, Fon M, Lee Y J, Phinney B S, Sadka A, Blumwald E. 2007. The citrus fruit proteome:insights into citrus fruit metabolism. Planta, 226:989-1005. pmid: 17541628
[34]	Ku H S, Pratt H K, Spurr A R, Harris W M. 1968. Isolation of active mitochondria from tomato fruit. Plant Physiology, 43 (6):883-887. doi: 10.1104/pp.43.6.883 pmid: 16656857
[35]	Kuhnert F, Stefanski A, Overbeck N, Drews L, Reichert A S, Stühler K, Weber A P M. 2020. Rapid single-step affinity purification of HA-tagged plant mitochondria. Plant Physiology, 182 (2):692-706. doi: 10.1104/pp.19.00732 pmid: 31818904
[36]	Lance C, Hobson G E, Young R E, Biale J B. 1965. Metabolic processes in cytoplasmic particles of the avocado fruit. VII. Oxidative and phosphorylative activities throughout the climacteric cycle. Plant Physiology, 40 (6):1116-1123. pmid: 5842698
[37]	Lange D L, Kader A A. 1997a. Changes in alternative pathway and mitochondrial respiration in avocado in response to elevated carbon dioxide levels. Journal of the American Society for Horticultural Science, 122 (2):245-252.
[38]	Lange D L, Kader A A. 1997b. Effects of elevated carbon dioxide on key mitochondrial respiratory enzymes in‘Hass’avocado fruit and fruit disks. Journal of the American Society for Horticultural Science, 122 (2):238-244.
[39]	Lázaro J J, Jiménez A, Camejo D, Iglesias-Baena I, del Carmen Martí M, Lázaro-Payo A, Barranco-Medina S, Sevilla F. 2013. Dissecting the integrative antioxidant and redox systems in plant mitochondria. Effect of stress and S-nitrosylation. Frontiers in Plant Science, 4:460. doi: 10.3389/fpls.2013.00460 pmid: 24348485
[40]	Li C, Cheng Y, Hou J, Zhu J, Sun L, Ge Y. 2021a. Application of methyl jasmonate postharvest maintains the quality of Nanguo pears by regulating mitochondrial energy metabolism. Journal of Integrative Agriculture, 20 (11):3075-3083.
[41]	Li P, Yin F, Song L, Zheng X. 2016. Alleviation of chilling injury in tomato fruit by exogenous application of oxalic acid. Food Chemistry, 202:125-132. doi: 10.1016/j.foodchem.2016.01.142 pmid: 26920276
[42]	Li S, Jiang H, Wang Y, Lyu L, Prusky D, Ji Y, Zheng X, Bi Y. 2020. Effect of benzothiadiazole treatment on improving the mitochondrial energy metabolism involved in induced resistance of apple fruit during postharvest storage. Food Chemistry, 302:125288.
[43]	Li Wanjie, Hu Kangdi. 2015. Common centrifugal techniques and their applications in laboratory. Bulletin of Biology, 50 (4):10-12. (in Chinese)
	李万杰, 胡康棣. 2015. 实验室常用离心技术与应用. 生物学通报, 50 (4):10-12.
[44]	Li X, Chai Y, Yang H, Tian Z, Li C, Xu R, Shi C, Zhu F, Zeng Y, Deng X, Wang P, Cheng Y. 2021b. Isolation and comparative proteomic analysis of mitochondria from the pulp of ripening citrus fruit. Horticulture Research, 8:31.
[45]	Li X, Tian Z, Chai Y, Yang H, Zhang M, Yang C, Xu R, Zhu F, Zeng Y, Deng X, Wang P, Cheng Y. 2022. Cytological and proteomic evidence reveals the involvement of mitochondria in hypoxia-induced quality degradation in postharvest citrus fruit. Food Chemistry, 375:131833.
[46]	Lieberman M. 1958. Isolation of cytoplasmic particles with cytochrome oxidase activity from apples. Science, 127 (3291):189-190. pmid: 17738487
[47]	Lieberman M. 1960. Oxidative activity of cytoplasmic particles of apples:electron transfer chain. Plant Physiology, 35 (6):796-801. doi: 10.1104/pp.35.6.796 pmid: 16655424
[48]	Liu Meijun, Sun Xuejuan, Zhang Zishan, Liu Yuting, Gao Huiyuan. 2014. Quantitative research of plant mitochondrial respiration state and its application in plant biology. Plant Physiology Journal, 50 (1):111-116. (in Chinese)
	刘美君, 孙学娟, 张子山, 刘玉亭, 高辉远. 2014. 植物线粒体呼吸状态的研究方法及其在植物生物学中的应用. 植物生理学报, 50 (1):111-116.
[49]	Liu S, Jing G, Zhu S. 2022. Nitric oxide(NO)involved in antioxidant enzyme gene regulation to delay mitochondrial damage in peach fruit. Postharvest Biology and Technology, 192:111993.
[50]	López-Vidal O, Camejo D, Rivera-Cabrera F, Konigsberg M, Villa-Hernández J M, Mendoza-Espinoza J A, Pérez-Flores L J, Sevilla F, Jiménez A, de León-Sánchez F D. 2016. Mitochondrial ascorbate-glutathione cycle and proteomic analysis of carbonylated proteins during tomato(Solanum lycopersicum)fruit ripening. Food Chemistry, 194:1064-1072. doi: 10.1016/j.foodchem.2015.08.055 pmid: 26471654
[51]	Luo L, He Y, Xu Q, Lyu W, Yan J, Xin P, Zhang D, Chu J, Li J, Yu H. 2020. Rapid and specific isolation of intact mitochondria from Arabidopsis leaves. Journal of Genetics and Genomics, 47 (1):65-68.
[52]	Lyu L, Bi Y, Li S, Xue H, Li Y, Prusky D B. 2019a. Sodium silicate prime defense responses in harvested muskmelon by regulating mitochondrial energy metabolism and reactive oxygen species production. Food Chemistry, 289:369-376.
[53]	Lyu L, Bi Y, Li S, Xue H, Zhang Z, Prusky D B. 2019b. Early defense responses involved in mitochondrial energy metabolism and reactive oxygen species accumulation in harvested muskmelons infected by Trichothecium roseum. Journal of Agricultural and Food Chemistry, 67 (15):4337-4345.
[54]	Miller L A, Romani R J. 1966. Sucrose density gradient distribution of mitochondrial protein and enzymes from preclimacteric and climacteric pears. Plant Physiology, 41 (3):411-414. doi: 10.1104/pp.41.3.411 pmid: 16656269
[55]	Moreau F, Romani R. 1982. Preparation of avocado mitochondria using self-generated Percoll density gradients and changes in buoyant density during ripening. Plant Physiology, 70 (5):1380-1384. doi: 10.1104/pp.70.5.1380 pmid: 16662683
[56]	Moriguchi T, Romani R J. 1995. Mitochondrial self-restoration as an index to the capacity of avocado fruit to sustain atmospheric stress at two climacteric states. Journal of the American Society for Horticultural Science, 120 (4):643-649.
[57]	Navet R, Jarmuszkiewicz W, Almeida A M, Sluse-Goffart C, Sluse F E. 2003. Energy conservation and dissipation in mitochondria isolated from developing tomato fruit of ethylene-defective mutants failing normal ripening: the effect of ethephon,a chemical precursor of ethylene. Journal of Bioenergetics and Biomembranes, 35:157-168.
[58]	Neal G E, Hulme A C. 1958. Activity of cytoplasmic particles isolated from apples. Nature, 181:1469-1470.
[59]	Niehaus M, Straube H, Künzler P, Rugen N, Hegermann J, Giavalisco P, Eubel H, Witte C P, Herde M. 2020. Rapid affinity purification of tagged plant mitochondria(Mito-AP)for metabolome and proteome analyses. Plant Physiology, 182 (3):1194-1210. doi: 10.1104/pp.19.00736 pmid: 31911558
[60]	Nukuntornprakit O, Luengwilai K, Siriphanich J. 2020. Chilling injury in pineapple fruit is related to mitochondrial antioxidative metabolism. Postharvest Biology and Technology, 170:111330.
[61]	Padwal-Desai S R, Ahmed E M, Dennison R A. 1969. Some properties of mitochondria from irradiated tomato fruit. Journal of Food Science, 34 (4):332-335.
[62]	Phelps D C, McDonald R E. 1990. Inhibition of electron transport activities in mitochondria from avocado and pepper fruit by naturally occurring polyamines. Physiologia Plantarum, 78 (1):15-21.
[63]	Piechowiak T. 2021. Effect of ozone-treatment on glutathione(GSH)status in selected berry fruit. Phytochemistry, 187:112767. doi: 10.1016/j.phytochem.2021.112767 pmid: 33838586
[64]	Piechowiak T, Sowa P, Balawejder M. 2021a. Effect of ozonation process on the energy metabolism in raspberry fruit during storage at room temperature. Food and Bioprocess Technology, 14:483-491.
[65]	Piechowiak T, Sowa P, Tarapatskyy M, Balawejder M. 2021b. The role of mitochondrial energy metabolism in shaping the quality of highbush blueberry fruit during storage in ozone enriched atmosphere. Food and Bioprocess Technology, 14:1973-1982.
[66]	Prasanna V, Prabha T N, Tharanathan R N. 2007. Fruit ripening phenomena-an overview. Critical Reviews in Food Science and Nutrition, 47 (1):1-19. doi: 10.1080/10408390600976841 pmid: 17364693
[67]	Qin G, Meng X, Wang Q, Tian S. 2009a. Oxidative damage of mitochondrial proteins contributes to fruit senescence:a redox proteomics analysis. Journal of Proteome Research, 8 (5):2449-2462.
[68]	Qin G, Wang Q, Liu J, Li B, Tian S. 2009b. Proteomic analysis of changes in mitochondrial protein expression during fruit senescence. Proteomics, 9 (17):4241-4253.
[69]	Raison J K, Lyons J M. 1970. The influence of mitochondrial concentration and storage on the respiratory control of isolated plant mitochondria. Plant Physiology, 45 (4):382-385. pmid: 5427107
[70]	Romani R J, Biale J B. 1957. Metabolic processes in cytoplasmic particles of the avocado fruit. IV. Ripening and the supernatant fraction. Plant Physiology, 32 (6):662-668. doi: 10.1104/pp.32.6.662 pmid: 16655063
[71]	Romani R J, Breidenbach R W, van Kooy J G. 1965. Isolation,yield,and fatty acid composition of intracellular particles from ripening fruits. Plant Physiology, 40 (3):561-566. doi: 10.1104/pp.40.3.561 pmid: 16656126
[72]	Romani R J, Ozelkok S. 1973. “Survival”of mitochondria in vitro:physical and energy parameters. Plant Physiology, 51 (4):702-707. doi: 10.1104/pp.51.4.702 pmid: 16658395
[73]	Romani R J, Tuskes S E, ÖzelkÖk S. 1974. Survival of plant mitochondria in vitro:form and function after 4 days at 25℃. Archives of Biochemistry and Biophysics, 164 (2):743-751. pmid: 4460886
[74]	Romani R J, Yu I K, Fisher L K. 1969. Isolation of tightly coupled mitochondria from acidic plant tissues. Plant Physiology, 44 (2):311-312. doi: 10.1104/pp.44.2.311 pmid: 16657062
[75]	Romani R J, Yu I K, Ku L L, Fisher L K, Dehgan N. 1968. Cellular senescence,radiation damage to mitochondria,and the compensatory response in ripening pear fruits. Plant Physiology, 43 (7):1089-1096. doi: 10.1104/pp.43.7.1089 pmid: 16656887
[76]	Shipway M R, Bramlage W J. 1973. Effects of carbon dioxide on activity of apple mitochondria. Plant Physiology, 51 (6):1095-1098. doi: 10.1104/pp.51.6.1095 pmid: 16658473
[77]	Tager J M. 1958. Isolation of active mitochondria from apples. Nature, 182:1521-1522.
[78]	Tian Shiping. 2013. Molecular mechanisms of fruit ripening and senescence. Chinese Bulletin of Botany, 48 (5):481-488. (in Chinese)
	田世平. 2013. 果实成熟和衰老的分子调控机制. 植物学报, 48 (5):481-488. doi: 10.3724/SP.J.1259.2013.00481
[79]	Vines H M, Oberbacher M F. 1965. Response of oxidation and phosphorylation in citrus mitochondria to arsenate. Nature, 206:319-320.
[80]	Wang C, Huang D, Tian W, Zhu S. 2021. Nitric oxide alleviates mitochondrial oxidative damage and maintains mitochondrial functions in peach fruit during cold storage. Scientia Horticulturae, 287:110249.
[81]	Wang Yuhang, Li Dou, Wang Chunheng, Jin Xin, Cheng Yajuan, Dai Zibo, Feng Lidan, Yang Jiangshan. 2024. The effect of melatonin on the subcellular reactive oxygen species metabolism during development and senescence in grape leaves. Acta Horticulturae Sinica, 51 (1):103-120. (in Chinese) doi: 10.16420/j.issn.0513-353x.2022-1214
	王宇航, 李斗, 王春恒, 金鑫, 陈亚娟, 戴子博, 冯丽丹, 杨江山. 2024. 褪黑素对葡萄叶片发育衰老过程中亚细胞活性氧代谢的影响. 园艺学报, 51 (1):103-120. doi: 10.16420/j.issn.0513-353x.2022-1214
[82]	Wiskich J T, Young R E, Biale J B. 1964. Metabolic processes in cytoplasmic particles of the avocado fruit. VI. Controlled oxidations and coupled phosphorylations. Plant Physiology, 39 (3):312-322. doi: 10.1104/pp.39.3.312 pmid: 16655919
[83]	Wu X, Jiang L, Yu M, An X, Ma R, Yu Z. 2016. Proteomic analysis of changes in mitochondrial protein expression during peach fruit ripening and senescence. Journal of Proteomics, 147:197-211. doi: S1874-3919(16)30244-5 pmid: 27288903
[84]	Yang Z, Cao S, Su X, Jiang Y. 2014. Respiratory activity and mitochondrial membrane associated with fruit senescence in postharvest peaches in response to UV-C treatment. Food Chemistry, 161:16-21. doi: 10.1016/j.foodchem.2014.03.120 pmid: 24837916
[85]	Yang Z, Zhu S, Wang X, Chen C, Huang D, Feng J. 2023. Nitric oxide modulates folate-mediated one-carbon metabolism and mitochondrial energy levels of peaches during cold storage. Frontiers in Nutrition, 10:1184736.
[86]	Zhou Q, Zhang C, Cheng S, Wei B, Liu X, Ji S. 2014. Changes in energy metabolism accompanying pitting in blueberries stored at low temperature. Food Chemistry, 164:493-501. doi: 10.1016/j.foodchem.2014.05.063 pmid: 24996362

种类 Category	果实 Fruit	组织 Tissue	分离方法 Method	参考文献 Reference
仁果类 Pome fruit	苹果 Apple	果肉Pulp	DC (1R)	Lieberman，1958
		果肉Pulp	DC (1R) + W (1T)	Neal & Hulme，1958；Tager，1958； Lieberman，1960
		果皮Peel	DC (1R) + W (1T)	Hulme et al.，1964
		果肉Pulp	DC (2R)	Romani et al.，1965
		果肉Pulp	DC (2R) + DPGC + SPGC + W (2T)	Qin et al.，2009b
	梨 Pear	果肉Pulp	DC (2R)	Romani et al.，1965
			DC (1R) + W (1T) + CSGC	Miller & Romani，1966
			DC (1R) + W (1T)	Li et al.，2021a
核果类 Stone fruit	鳄梨 Avocado	果肉Pulp	DC (1R) + W (1T)	Biale et al.，1957
			DC (3R)	Wiskich et al.，1964
			DC (2R)	Lance et al.，1965；Hobson et al.，1966
			DC (1R) + W (1T) + SPGC + W (2T)	Moreau & Romani，1982
	芒果 Mango	果肉Pulp	DC (2R)	Baqui et al.，1974
	芒果 Mango	果肉Pulp	DC (1R) + W (1T)	Kane & Marcellin，1978
	桃 Peach	果肉Pulp	DC (1R) + W (1T)	Kan et al.，2010，2011
			DC (1R) + W (2T)	Jin et al.，2013
			DC (1R) + DPGC + W (1T) + SPGC + W (2T)	Qin et al.，2009a
			DC (1R) + DPGC + W (2T)	Yang et al.，2014
			DC (1R) + W (1T) + DSGC + W (1T)	Jing et al.，2016
			DC (1R) + DPGC + W (2T)	Wu et al.，2016
			DC (1R) + DSGC + W (1T)	Wang et al.，2021
浆果类 Berries	番茄 Tomato	果肉Pulp	DC (1R) + W (1T)	Dickinson & Hanson，1965；Dickinson et al.，1970；Costa et al.，1999
			DC (2R)	Ku et al.，1968；Raison & Lyons，1970
			DC (1R) + W (2T)	Li et al.，2016
			DC (1R) + W (1T) + CSGC	Kalra & Brooks，1973
			DC (2R) + DPGC	Jeffery et al.，1986
			DC (1R) + W (2T) + SPGC + W (2T)	Jarmuszkiewicz et al.，1998
			DC (1R) + SPGC + W (2T)	Holtzapffel et al.，2002
			DC (2R) + DPGC + W (2T)	de Oliveira et al.，2017
			DC (1R) + DPGC + W (2T) + SPGC + W (2T)	Cai et al.，2018
	香蕉 Banana	果肉Pulp	DC (2R)	Haard & Hultin，1968
	香蕉 Banana	果皮Peel	DC (1R) + W (2T) + CSGC + W (1T)	Chotikakham et al.，2022
	蓝莓 Blueberry	果肉Pulp	DC (1R) + W (1T)	Zhou et al.，2014；Piechowiak，2021
	番木瓜Papaya 草莓Strawberry 番石榴Guava	果肉Pulp	DC (2R) + DPGC + W (2T)	de Oliveira et al.，2017
	猕猴桃Kiwifruit	果肉Pulp	DC (1R) + W (2T)	Ali et al.，2020
	菠萝 Pineapple	果肉Pulp	DC (1R) + DSGC	Nukuntornprakit et al.，2020
柑果类 Citrus	柠檬Lemon	果肉Pulp	DC (1R) + W (1T)	Bogin & Erickson，1965
	甜橙Orange		DC (2R)	Vines & Oberbacher，1965
	甜橙Orange		DC (1R)	Katz et al.，2007
	柑Tangerine 橘Mandarin 橙Orange 柚Pummelo		DC (2R) + DPGC + W (2T)	Li et al.，2021b
瓜类 Melon	甜瓜 Muskmelon	果肉Pulp	DC (2R) + DPGC + SPGC + W (2T)	Lyu et al.，2019a，2019b

种类 Category	果实 Fruit	组织 Tissue	分离方法 Method	参考文献 Reference
仁果类 Pome fruit	苹果 Apple	果肉Pulp	DC (1R)	Lieberman，1958
		果肉Pulp	DC (1R) + W (1T)	Neal & Hulme，1958；Tager，1958； Lieberman，1960
		果皮Peel	DC (1R) + W (1T)	Hulme et al.，1964
		果肉Pulp	DC (2R)	Romani et al.，1965
		果肉Pulp	DC (2R) + DPGC + SPGC + W (2T)	Qin et al.，2009b
	梨 Pear	果肉Pulp	DC (2R)	Romani et al.，1965
			DC (1R) + W (1T) + CSGC	Miller & Romani，1966
			DC (1R) + W (1T)	Li et al.，2021a
核果类 Stone fruit	鳄梨 Avocado	果肉Pulp	DC (1R) + W (1T)	Biale et al.，1957
			DC (3R)	Wiskich et al.，1964
			DC (2R)	Lance et al.，1965；Hobson et al.，1966
			DC (1R) + W (1T) + SPGC + W (2T)	Moreau & Romani，1982
	芒果 Mango	果肉Pulp	DC (2R)	Baqui et al.，1974
	芒果 Mango	果肉Pulp	DC (1R) + W (1T)	Kane & Marcellin，1978
	桃 Peach	果肉Pulp	DC (1R) + W (1T)	Kan et al.，2010，2011
			DC (1R) + W (2T)	Jin et al.，2013
			DC (1R) + DPGC + W (1T) + SPGC + W (2T)	Qin et al.，2009a
			DC (1R) + DPGC + W (2T)	Yang et al.，2014
			DC (1R) + W (1T) + DSGC + W (1T)	Jing et al.，2016
			DC (1R) + DPGC + W (2T)	Wu et al.，2016
			DC (1R) + DSGC + W (1T)	Wang et al.，2021
浆果类 Berries	番茄 Tomato	果肉Pulp	DC (1R) + W (1T)	Dickinson & Hanson，1965；Dickinson et al.，1970；Costa et al.，1999
			DC (2R)	Ku et al.，1968；Raison & Lyons，1970
			DC (1R) + W (2T)	Li et al.，2016
			DC (1R) + W (1T) + CSGC	Kalra & Brooks，1973
			DC (2R) + DPGC	Jeffery et al.，1986
			DC (1R) + W (2T) + SPGC + W (2T)	Jarmuszkiewicz et al.，1998
			DC (1R) + SPGC + W (2T)	Holtzapffel et al.，2002
			DC (2R) + DPGC + W (2T)	de Oliveira et al.，2017
			DC (1R) + DPGC + W (2T) + SPGC + W (2T)	Cai et al.，2018
	香蕉 Banana	果肉Pulp	DC (2R)	Haard & Hultin，1968
	香蕉 Banana	果皮Peel	DC (1R) + W (2T) + CSGC + W (1T)	Chotikakham et al.，2022
	蓝莓 Blueberry	果肉Pulp	DC (1R) + W (1T)	Zhou et al.，2014；Piechowiak，2021
	番木瓜Papaya 草莓Strawberry 番石榴Guava	果肉Pulp	DC (2R) + DPGC + W (2T)	de Oliveira et al.，2017
	猕猴桃Kiwifruit	果肉Pulp	DC (1R) + W (2T)	Ali et al.，2020
	菠萝 Pineapple	果肉Pulp	DC (1R) + DSGC	Nukuntornprakit et al.，2020
柑果类 Citrus	柠檬Lemon	果肉Pulp	DC (1R) + W (1T)	Bogin & Erickson，1965
	甜橙Orange		DC (2R)	Vines & Oberbacher，1965
	甜橙Orange		DC (1R)	Katz et al.，2007
	柑Tangerine 橘Mandarin 橙Orange 柚Pummelo		DC (2R) + DPGC + W (2T)	Li et al.，2021b
瓜类 Melon	甜瓜 Muskmelon	果肉Pulp	DC (2R) + DPGC + SPGC + W (2T)	Lyu et al.，2019a，2019b

物质类别 Substrate categories	代表物质及文献中报道的浓度（范围） Components and concentration	作用及备注 Function and note
缓冲物质 Buffer material	磷酸盐（Phosphate，0.01 ~ 0.50 mol · L^-1），Tris（0.02 ~ 0.50 mol · L^-1），Na-barbital（0.05 mol · L^-1），Na₄P₂O₇（0.025 mol · L^-1），MOPS（0.05 ~ 0.50 mol · L^-1），TES（0.025 mol · L^-1），Bicine（0.1 mol · L^-1），Hepes（0.01 ~ 0.04 mol · L^-1）	维持匀浆液pH稳定；根据果实样品特性调整缓冲物质的浓度；可同时使用多种缓冲物质 Keeping the pH of homogenate stable；Adjusting the concentration of buffer material according to the characteristics of the fruit samples；Multiple buffer materials can be used simultaneously
渗透压调节物质 Osmolyte	蔗糖（Sucrose，0.15 ~ 0.70 mol · L^-1），甘露醇（Mannitol）/山梨醇（Sorbitol）（0.25 ~ 0.60 mol · L^-1）	调节溶液渗透压以维持线粒体膜结构稳定；可同时使用多种渗透压调节物质 Regulation of solution osmotic pressure to maintain the stability of mitochondrial membrane structure；Multiple osmolytes can be used simultaneously
抗氧化物质 Antioxidant	抗坏血酸（Ascorbic acid，0.01 ~ 0.03 mol · L^-1），柠檬酸（Citric acid，0.02 mol · L^-1），半胱氨酸（Cysteine，1 ~ 10 mmol · L^-1或0.05% ~ 0.10% [w/v]），DTT（0.5 ~ 2.0 mmol · L^-1），β-巯基乙醇（β-Mercaptoethanol，5 ~ 10 mmol L^-1或0.1% [v/v]）	维持多酚的还原性状态，延缓匀浆过程中的褐变；保护酶分子的还原性基团，稳定酶活性；用时现加 Maintaining the reducing state of polyphenols and delaying browning during homogenization；Protecting the reducing groups on the enzyme molecule and keeping the enzyme activity stable；Adding as needed
螯合剂 Chelator	EDTA（0.2 ~ 10.0 mmol · L^-1），EGTA（0.2 ~ 1.0 mmol · L^-1）	去除重金属离子对酶的抑制作用；浓度高时会抑制酶活性 Removing the inhibitory effect of heavy metal ions on enzymes；Inhibiting the enzyme activity at high concentrations
其他 Other	PVP（0.05% ~ 5.00% [w/v]），PVPP（1.5% [w/v]）	结合多酚以降低其对线粒体酶活性的抑制 Binding polyphenols to reduce the inhibition on mitochondrial enzyme activity
	BSA（0.05% ~ 1.00% [w/v]）	降低脂肪酸对线粒体活性的抑制 Reducing the inhibition of fatty acids on mitochondrial enzyme activity
	MgCl₂（1 ~ 6 mmol · L^-1），KCl（6 ~ 10 mmol · L^-1），MgSO₄（2 mmol · L^-1）	低浓度时可降低线粒体聚集且有利于酶活性，但高浓度时会加剧线粒体聚集 Reducing mitochondrial aggregation and promoting the enzyme activity at low concentrations；Increasing mitochondrial aggregation at high concentrations

物质类别 Substrate categories	代表物质及文献中报道的浓度（范围） Components and concentration	作用及备注 Function and note
缓冲物质 Buffer material	磷酸盐（Phosphate，0.01 ~ 0.50 mol · L^-1），Tris（0.02 ~ 0.50 mol · L^-1），Na-barbital（0.05 mol · L^-1），Na₄P₂O₇（0.025 mol · L^-1），MOPS（0.05 ~ 0.50 mol · L^-1），TES（0.025 mol · L^-1），Bicine（0.1 mol · L^-1），Hepes（0.01 ~ 0.04 mol · L^-1）	维持匀浆液pH稳定；根据果实样品特性调整缓冲物质的浓度；可同时使用多种缓冲物质 Keeping the pH of homogenate stable；Adjusting the concentration of buffer material according to the characteristics of the fruit samples；Multiple buffer materials can be used simultaneously
渗透压调节物质 Osmolyte	蔗糖（Sucrose，0.15 ~ 0.70 mol · L^-1），甘露醇（Mannitol）/山梨醇（Sorbitol）（0.25 ~ 0.60 mol · L^-1）	调节溶液渗透压以维持线粒体膜结构稳定；可同时使用多种渗透压调节物质 Regulation of solution osmotic pressure to maintain the stability of mitochondrial membrane structure；Multiple osmolytes can be used simultaneously
抗氧化物质 Antioxidant	抗坏血酸（Ascorbic acid，0.01 ~ 0.03 mol · L^-1），柠檬酸（Citric acid，0.02 mol · L^-1），半胱氨酸（Cysteine，1 ~ 10 mmol · L^-1或0.05% ~ 0.10% [w/v]），DTT（0.5 ~ 2.0 mmol · L^-1），β-巯基乙醇（β-Mercaptoethanol，5 ~ 10 mmol L^-1或0.1% [v/v]）	维持多酚的还原性状态，延缓匀浆过程中的褐变；保护酶分子的还原性基团，稳定酶活性；用时现加 Maintaining the reducing state of polyphenols and delaying browning during homogenization；Protecting the reducing groups on the enzyme molecule and keeping the enzyme activity stable；Adding as needed
螯合剂 Chelator	EDTA（0.2 ~ 10.0 mmol · L^-1），EGTA（0.2 ~ 1.0 mmol · L^-1）	去除重金属离子对酶的抑制作用；浓度高时会抑制酶活性 Removing the inhibitory effect of heavy metal ions on enzymes；Inhibiting the enzyme activity at high concentrations
其他 Other	PVP（0.05% ~ 5.00% [w/v]），PVPP（1.5% [w/v]）	结合多酚以降低其对线粒体酶活性的抑制 Binding polyphenols to reduce the inhibition on mitochondrial enzyme activity
	BSA（0.05% ~ 1.00% [w/v]）	降低脂肪酸对线粒体活性的抑制 Reducing the inhibition of fatty acids on mitochondrial enzyme activity
	MgCl₂（1 ~ 6 mmol · L^-1），KCl（6 ~ 10 mmol · L^-1），MgSO₄（2 mmol · L^-1）	低浓度时可降低线粒体聚集且有利于酶活性，但高浓度时会加剧线粒体聚集 Reducing mitochondrial aggregation and promoting the enzyme activity at low concentrations；Increasing mitochondrial aggregation at high concentrations