Soil dehydrogenase determination method.

asked: Which man has done the soil
dhydrogenase
activity of the experiment, please point to the mystery ah, thank you first. When I do the standard curve, after adding the extractor (methanol or acetone), the upper part of each tube appears precipitation, the red solution is located in the lower part of the tube, upside down after shaking the tubes are cloudy. After a period of time each tube is clarified, but are colorless, if not upside down and just oscillating will eventually become a clarification solution of color, why ah?
production of the standard curve
1.1 preparation of different concentrations of TTC liquid: 1,2,3,4,5,6,7ml from 1 mg/ml TTC solution, In a 50 ml bottle, the contents of TTC were reduced to 50 ml by distilled water, and the concentration of TTC in each bottle was 20, 40, 60, 80., 100, 120, 140ug/ml.
1.2 Add tris-hydrochloric acid buffer (pH=8.4) to each plug-in
centrifugal tube
(pH=8.4) 2 ml plus 2 ml distilled water plus 1 ml TTC liquid (from low to high concentrations in turn) ), the control tube is added with 2 ml tris-hydrochloric acid buffer and 3 ml distilled water, without TTC, the resulting TTC content of each centrifuge tube is 20, 40, 60, 80., 100, 120, 140ug, respectively.
1.3 Each tube is added with sodium dithionate 1 g, oscillating and shaking well, so that the TTC is reduced to produce a red TPF.
1.4 to add 5 ml of methanol to each tube, oscillating extraction 3min, extraction of TPF.
1.5 to take the liquid at 485 nm wavelength to measure the absorbent value.
1.7 to plot the standard curve.
: I have done dehydrogenase activity determination, the basic steps are the same, but the final use of toluene extraction, made the standard curve is good, but there is a colorless phenomenon like above, the appearance of colorless will not be due to the extraction of the problem.
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