Lammertyn J, Franck C, Verlinden B E, Nicolaï B M
Flanders Centre/Laboratorium of Postharvest Technology, Katholieke Universiteit Leuven, Willem de Croylaan 42, B-3001, Leuven, Belgium.
J Exp Bot. 2001 Sep;52(362):1769-77. doi: 10.1093/jexbot/52.362.1769.
The influence of the O(2) and CO(2) concentration and the temperature on the O(2) uptake rate of cool-stored intact pears and pear cell protoplasts in suspension was compared. Protocols to isolate pear cell protoplasts from pear tissue and two methods to measure protoplast respiration have been developed. Modified Michaelis-Menten kinetics were applied to describe the effect of the O(2) and the CO(2) concentration on the O(2) uptake rate and temperature dependence was analysed with an Arrhenius equation. Both systems were described with a non-competitive type of CO(2) inhibition. Due to the inclusion of gas diffusion properties, the Michaelis-Menten constant for intact pears (2.5 mM) was significantly larger than the one for protoplasts in suspension (3 microM), which was in turn larger than the Michaelis-Menten constant obtained in mitochondrial respiration measurements described in the literature. It was calculated that only 3.6% of the total diffusion effect absorbed in the Michaelis-Menten constant for intact pears, could be attributed to intracellular gas diffusion. The number of cells per volume of tissue was counted microscopically to establish a relationship between the pear cell protoplast and intact pear O(2) uptake rate. A remarkable similarity was observed: values of 61.8 nmol kg(-1) s(-1) for protoplasts and 87.1 nmol kg(-1) s(-1) for intact pears were obtained. Also, the inhibitory effect of CO(2) on the respiration rate was almost identical for protoplasts and intact pears, suggesting that protoplast suspensions are useful for the study of other aspects of the respiration metabolism.
比较了氧气（O₂）和二氧化碳（CO₂）浓度以及温度对冷藏完整梨和悬浮梨细胞原生质体氧气吸收速率的影响。已开发出从梨组织中分离梨细胞原生质体的方案以及两种测量原生质体呼吸作用的方法。应用修正的米氏动力学来描述氧气和二氧化碳浓度对氧气吸收速率的影响，并使用阿伦尼乌斯方程分析温度依赖性。两个系统均表现出非竞争性的二氧化碳抑制类型。由于考虑了气体扩散特性，完整梨的米氏常数（2.5 mM）显著大于悬浮原生质体的米氏常数（3 μM），而悬浮原生质体的米氏常数又大于文献中描述的线粒体呼吸测量所得的米氏常数。经计算，完整梨米氏常数中吸收的总扩散效应中只有3.6%可归因于细胞内气体扩散。通过显微镜计数单位体积组织中的细胞数量，以建立梨细胞原生质体与完整梨氧气吸收速率之间的关系。观察到显著的相似性：原生质体的氧气吸收速率值为61.8 nmol kg⁻¹ s⁻¹，完整梨的氧气吸收速率值为87.1 nmol kg⁻¹ s⁻¹。此外，二氧化碳对原生质体和完整梨呼吸速率的抑制作用几乎相同，这表明原生质体悬浮液可用于呼吸代谢其他方面的研究。
