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After the reaction, sample was extracted by petroleum ether with pH adjustment and dried under vacuum.
A standard reaction sample was composed of a mixture containing enzyme and 0.5% (w/v) β-glucan incubated in 10 mM NaPO4 buffer, pH 7.0.
Following a 4 hr incubation, the reaction sample was extracted by LiCl method, then analyzed by ethidium bromide staining/UV illumination and autoradiography.
50 µl of the reaction sample was added to 600 µl of peroxoquant reagent and incubated for 30 minutes and the samples were read at 560 nm.
At designated incubation intervals (5, 10 & 15 minutes), 75 µl of the reaction sample was taken and quenched with 25 µl of acetonitrile containing the internal standard and centrifuged.
Equal amounts of cDNA were used in each reaction sample.
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Reaction samples were withdrawn at regular intervals, and OD405 was measured immediately using a microplate reader.
The reaction samples were withdrawn at regular intervals for 4 h and analyzed by HPLC.
The ginsenosides in the reaction samples obtained were identified as the same retention times of the ginsenoside standards.
Liquid reaction samples were analyzed offline using gas chromatography coupled with mass spectrometer (Shimadzu QP5050) using HP-5 column.
After the treatment period, to arrest the reaction samples were diluted immediately with sterile phosphate buffer (50 mM, pH 7).
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