Künnecke B, Seelig J
Biocenter, University of Basel, Switzerland.
Biochim Biophys Acta. 1991 Oct 26;1095(2):103-13. doi: 10.1016/0167-4889(91)90071-5.
The metabolism of glucose to glycogen in the liver of fasted and well-fed rats was investigated with 13C nuclear magnetic resonance spectroscopy using [1,2-(13)C2]glucose as the main substrate. The unique spectroscopic feature of this molecule is the 13C-13C homonuclear coupling leading to characteristic doublets for the C-1 and C-2 resonances of glucose and its breakdown products as long as the two 13C nuclei remain bonded together. The doublet resonances of [1,2-(13)C2]glucose thus provide an ideal marker to follow the fate of this exogenous substrate through the metabolic pathways. [1,2-(13)C2]Glucose was injected intraperitoneally into anesthetized rats and the in vivo 13C-NMR measurements of the intact animals revealed the transformation of the injected glucose into liver glycogen. Glycogen was extracted from the liver and high resolution 13C-NMR spectra were obtained before and after hydrolysis of glycogen. Intact [1,2-13C2]glucose molecules give rise to doublet resonances, natural abundance [13C]glucose molecules produce singlet resonances. From an analysis of the doublet-to-singlet intensities the following conclusions were derived. (i) In fasted rats virtually 100% of the glycosyl units in glycogen were 13C-NMR visible. In contrast, the 13C-NMR visibility of glycogen decreased to 30-40% in well-fed rats. (ii) In fed rats a minimum of 67 +/- 7% of the exogenous [1,2-(13)C2]glucose was incorporated into the liver glycogen via the direct pathway. No contribution of the indirect pathway could be detected. (iii) In fasted rats externally supplied glucose appeared to be consumed in different metabolic processes and less [1,2-(13)C2]glucose was found to be incorporated into glycogen (13 +/- 1%). However, the observation of [5,6-(13)C2]glucose in liver glycogen provided evidence for the operation of the so-called indirect pathway of glycogen synthesis. The activity of the indirect pathway was at least 9% but not more than 30% of the direct pathway. (vi) The pentose phosphate pathway was of little significance for glucose but became detectable upon injection of [1-(13)C]ribose.
采用[1,2-(13)C2]葡萄糖作为主要底物，运用13C核磁共振波谱技术研究了禁食和饱食大鼠肝脏中葡萄糖向糖原的代谢过程。该分子独特的波谱特征是13C-13C同核耦合，只要两个13C核保持结合在一起，就会导致葡萄糖及其分解产物的C-1和C-2共振出现特征性双峰。因此，[1,2-(13)C2]葡萄糖的双峰共振为追踪这种外源性底物在代谢途径中的命运提供了理想的标记。将[1,2-(13)C2]葡萄糖腹腔注射到麻醉的大鼠体内，对完整动物进行的体内13C-NMR测量揭示了注射的葡萄糖向肝糖原的转化。从肝脏中提取糖原，并在糖原水解前后获得高分辨率13C-NMR光谱。完整的[1,2-13C2]葡萄糖分子产生双峰共振，天然丰度的[13C]葡萄糖分子产生单峰共振。通过对双峰与单峰强度的分析得出了以下结论。(i)在禁食大鼠中，糖原中几乎100%的糖基单位在13C-NMR下可见。相比之下，在饱食大鼠中，糖原的13C-NMR可见性降至30-40%。(ii)在喂食大鼠中，至少67±7%的外源性[1,2-(13)C2]葡萄糖通过直接途径掺入肝糖原。未检测到间接途径的贡献。(iii)在禁食大鼠中，外源供应的葡萄糖似乎在不同的代谢过程中被消耗，发现较少的[1,2-(13)C2]葡萄糖掺入糖原(13±1%)。然而，在肝糖原中观察到[5,6-(13)C2]葡萄糖为糖原合成的所谓间接途径的运作提供了证据。间接途径的活性至少为直接途径的9%但不超过30%。(iv)磷酸戊糖途径对葡萄糖意义不大，但在注射[1-(13)C]核糖后可检测到。
