"Methylotrophic biomass as 2H-labeled substrate for biosynthesis of inosine"
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from MetOH (nd20 = 1.33). The purity of the product was judged by using controls of normal nucleosides, and running mixed TLC with graded amounts of the neighboring nucleosides. Quantitative determination During the growth inosine was separated by TLC on Silufol UV-254 plates with mobile phases: n -ButOH - AcOH - water (2:1:1, v/v) using pure commercial available inosine as a standard.
The amount of inosine was determined for 10 ml aliquots of liquid growth medium by TLC. The sports were eluted by 0.1 N solution of HCl (10 ml). The absorbance of the eluates was measured at 249 nm and the content of inosine was determined using a standard curve. The convertion of glucose was estimated enzymatically with glucoseoxydenase method . Equipment Absorbance was measured with a spectrophotometer
Beckman DU-6 (USA). The analysis of protein hydrolisates was carried out using a Biotronic LC 50001 chromatograph (Germany), 230 x 3.2 mm, working pressure 50-60 atm, flow-rate 18.5 ml/h. The levels of deuterium enrichment of amino acids were investigated with the aid of EI MS after derivatization to methyl esters of N-Dns-amino acids . FAB MS was performed on Hitachi MBA spectrometer (Japan) on glyserol template at potential 5 кV and
an ion current of 0.6-0.8 мА. RESULTS AND DISCUSSION Production of 2H-labeled inosine For biosynthesis of 2H- labeled inosine we employed bacterium Bacillus subtillis, which could produce and accumulate a conciderable amount of inosine exogeniously due to an altered nucleoside metabolism. This strain displayed the maximum productivity on FM medium, containing as a source of carbon and energy glucose (12 m/m.%), and as a source of growthСкачать сочинение