Appl Biochem Biotechnol. 2014 Jan;172(1):289-97. doi: 10.1007/s12010-013-0543-1.
In this study, an enzymatic procedure for the determination of glycine (Gly) was developed by using a column containing immobilized glutamate dehydrogenase (GDH) on glyoxal agarose beads. Ammonia is produced from the enzymatic reactions between Gly and GDH with NAD(+) in phosphate buffer medium. The indophenol blue method was used for ammonia detection based on the spectrophotometric measurements of blue-colored product absorbing at 640 nm. The calibration graph is linear in the range of 0.1-10 mM of Gly concentrations. The effect of pH, temperature, and time interval was studied to find column stability, and also the interference effects of other amino acids was investigated. The interaction between GDH and glyoxal agarose beads was analyzed by Fourier transform infrared (FTIR) spectroscopy. The morphology of the immobilized and non-immobilized agarose beads were characterized by atomic force microscopy (AFM).
在这项研究中，开发了一种使用含有固定化谷氨酸脱氢酶 (GDH) 的珠状乙二醛琼脂糖柱的酶法来测定甘氨酸 (Gly) 的方法。在磷酸盐缓冲介质中，GDH 与 Gly 和 NAD(+) 之间的酶促反应产生氨。根据分光光度法测量蓝色产物在 640nm 处的吸收，使用吲哚酚蓝法进行氨检测。校准曲线在 Gly 浓度为 0.1-10mM 的范围内呈线性。研究了 pH 值、温度和时间间隔对柱稳定性的影响，并研究了其他氨基酸的干扰效应。通过傅里叶变换红外 (FTIR) 光谱分析了 GDH 和乙二醛琼脂糖珠之间的相互作用。通过原子力显微镜 (AFM) 对固定化和非固定化琼脂糖珠的形态进行了表征。
