Department of Agronomy, Ohio State University, Wooster, Ohio 44691.
Plant Physiol. 1987 Nov;85(3):768-73. doi: 10.1104/pp.85.3.768.
Metabolites in Bradyrhizobium japonicum bacteroids and in Glycine max (L.) Merr. cytosol from root nodules were analyzed using an isolation technique which makes it possible to estimate and correct for changes in concentration which may occur during bacteroid isolation. Bacteroid and cytosol extracts were fractionated on ion-exchange columns and were analyzed for carbohydrate composition using gas-liquid chromatography and for organic acid and amino acid composition using high performance liquid chromatography. Analysis of organic acids in plant tissues as the phenacyl derivatives is reported for the first time and this approach revealed the presence of several unknown organic acids in nodules. The time required for separation of bacteroids and cytosol was varied, and significant change in concentration of individual compounds during the separation of the two fractions was estimated by calculating the regression of concentration on time. When a statistically significant slope was found, the true concentration was estimated by extrapolating the regression line to time zero. Of 78 concentration estimates made, there was a statistically significant (5% level) change in concentration during sample preparation for only five metabolites: glucose, sucrose, and succinate in the cytosol and d-pinitol and serine in bacteroids. On a mass basis, the major compounds in bacteroids were (descending order of concentration): myo-inositol, d-chiro-inositol, alpha,alpha-trehalose, sucrose, aspartate, glutamate, d-pinitol, arginine, malonate, and glucose. On a proportional basis (concentration in bacteroid as percent of concentration in bacteroid + cytosol fractions), the major compounds were: alpha-aminoadipate (94), trehalose (66), lysine (58), and arginine (46). The results indicate that metabolite concentrations in bacteroids can be reliably determined.
根瘤中慢生根瘤菌类菌体和大豆细胞质体的代谢物是使用一种分离技术进行分析的，这种技术可以对可能在类菌体分离过程中发生的浓度变化进行估计和修正。类菌体和细胞质体提取物在离子交换柱上进行分级，并使用气相色谱法分析碳水化合物组成，使用高效液相色谱法分析有机酸和氨基酸组成。首次报道了植物组织中有机酸作为苯甲酰衍生物的分析方法，这种方法揭示了在根瘤中存在几种未知的有机酸。类菌体和细胞质体的分离时间不同，通过计算浓度随时间的回归来估计两个分数分离过程中个别化合物浓度的显著变化。当发现统计上显著的斜率时，通过将回归线外推到时间零来估计真实浓度。在进行的 78 次浓度估计中，只有 5 种代谢物（细胞质体中的葡萄糖、蔗糖和琥珀酸，以及类菌体中的 D-松醇和丝氨酸）在样品制备过程中浓度发生了统计学上显著（5%水平）的变化。以质量为基础，类菌体中的主要化合物（按浓度递减顺序）为：肌醇、D-手性肌醇、α、α-海藻糖、蔗糖、天冬氨酸、谷氨酸、D-松醇、精氨酸、丙二酸和葡萄糖。以比例为基础（类菌体中浓度占类菌体+细胞质体分数的浓度），主要化合物为：α-氨基己二酸（94）、海藻糖（66）、赖氨酸（58）和精氨酸（46）。结果表明，可以可靠地确定类菌体中代谢物的浓度。
