ICL (ec4.1.3.1) mainly exists in plants and microorganisms. During the germination process of oil seeds,
fat is converted into carbohydrate through glyoxylate cycle and other processes. ICL is one of the key
enzymes in glyoxylate cycle.
The degradation of isocitric acid to glyoxylic acid and succinic acid is catalyzed by ICL. Glyoxylic acid
and NADH generate ethanol and NAD under the action of LDH. NADH has a characteristic absorption
peak at 340nm. Monitoring the reduction rate of 340nm absorbance can indirectly reflect the activity of
ICL.
Reagents and Equipment Required but Not Provided:
Spectrophotometer, water bath, desk centrifuge, adjustable transferpettor, 1mL quartz cuvette, mortar /
homogenizer, ice and distilled water.
Sample preparation:
1. Cells or bacteria: Collect bacteria or cells into centrifuge tube, after centrifugation discard supernatant.
Suggest 2 million of bacteria or cells with 0.4 mL of extract solution, splitting with ultrasonication (ice
bath, power 20%, work time 3s, interval 10s, for 30 times). Centrifuge at 15000 ×g for 10 minutes at 4℃