Wisnewski N, Saz H J, Mössinger J, deBruyn B S, Weinstein P P
Department of Biological Sciences, University of Notre Dame, Indiana 46556.
J Parasitol. 1990 Jun;76(3):302-6.
Radioisotopically labeled glucose and pyruvate were employed to elucidate biochemical mechanisms utilized by the filariid Dipetalonema viteae during cultivation. Adults isolated from amicrofilaremic hamsters were incubated at 37 C in a mixture of NCTC135:IMDM (NI), with either D-[14C-(U)]glucose or [1-14C]pyruvate, under a gas phase of 5% CO2/N2 for 3 days. Labeled organic acids were separated and quantified by ion exchange chromatography. High performance liquid chromatography (HPLC) was used for separation and quantification of the 23 free amino acids in the NI medium. Ion exchange chromatography revealed that lactate was the major glycolytic end product, accounting for 90-97% of the original carbon utilized. Small amounts of radioactivity were recovered in succinate and variably in acetate fractions. HPLC analysis demonstrated that some amino acids increased, some decreased, and some remained at the initial concentration. Alanine exhibited the greatest change, consistently increasing from 2 to 4 times the original concentration. Analyses of purified amino acid peaks revealed radioactivity only in the alanine peak, accounting for 2-4% of the original carbon utilized.
利用放射性同位素标记的葡萄糖和丙酮酸来阐明丝状线虫双瓣尾丝虫在培养过程中所利用的生化机制。从无微丝蚴血症的仓鼠体内分离出的成虫，于37℃在NCTC135:IMDM（NI）混合液中培养，分别加入D-[14C-(U)]葡萄糖或[1-14C]丙酮酸，在5% CO2/N2气相条件下培养3天。通过离子交换色谱法分离并定量标记的有机酸。采用高效液相色谱法（HPLC）分离并定量NI培养基中的23种游离氨基酸。离子交换色谱分析显示，乳酸是糖酵解的主要终产物，占所利用原始碳的90 - 97%。在琥珀酸中回收了少量放射性，乙酸盐组分中的放射性则变化不定。HPLC分析表明，一些氨基酸含量增加，一些减少，还有一些保持在初始浓度。丙氨酸变化最大，浓度始终从初始浓度增加到2至4倍。对纯化的氨基酸峰进行分析，结果显示仅在丙氨酸峰中检测到放射性，占所利用原始碳的2 - 4%。
