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(70,135,221); "> "experimental purpose"
to understand the production site of ketones and how to determine the production and utilization of ketones.
in liver mitochondrials, fatty acids are produced by β-oxidation of excess acetyl coenzyme A synthesized ketones. Ketones include acetylacetic acid, β-hydroxybutyric acid, and acetone,compounds . The liver cannot use ketones, only in the livertissue, especially in the heart and skeletal muscle, ketones can be converted into acetyl coenzyme A and oxidized use.
this experiment took butyric acid as the substitut, insulation with liver homogenizer, and then the production of ketones in liver plasma was determined. In addition, in the case of liver and muscle tissue coexist, the amount of ketone production is then measured. Under these two different conditions, we can understand the above theory by the difference of ketone body content. This experiment mainly measured the content of acetone.
principle of ketone body determination: iodine in alkaline solution can oxidize acetone into iodine imitation. By titring the remaining iodine with sodium sulfate, the iodine consumed can be calculated, and the content of ketones (represented by acetone) can also be calculated. The reaction is as follows:
1. Experimental equipment
"> test tube; piped tube; conical bottle; " and rack.
2. Experiment reagent
(1) 0.1% starch solution.
(2) 0.9% NaCl solution.
(3) 15% tricloste.
(4) 10% NaOH solution.
(5)10% HCl solution.
(6) 0.5mol/L butyric acid solution: take 5 ml butyric acid dissolved in 100 ml 0.5mol/L NaOH.
(7) 0.1mol/L iodine: I2 12.5g and KI 25g water dissolved, diluted to scale 1L, calibration with 0.1mol/L Na2S2O3.
(8) 0.02mol/L Na2S2O3: 24.82g Na2S2O3.5H2O and 400mg waterless Na2CO3 dissolved in 1L boiled water, with 0.1mol/L solution, labeled with 0.1mol/L KIO3. Dilute the calibration Na2S2O3 solution to 0.02mol/L when in use.(Experimental Operations)
1. Preparation of specimens:
kills the rabbit, removes the liver, and uses 0.9% Na Cl washed the stained blood, putfilter paper on the surface of the water, said to take liver tissue 5g in the research, plus a little 0.9% NaCl to the total volume of 10 ml, to make liver tissue homogeneity. In addition, the hind leg muscle 5g, according to the above method and proportion, to make muscle tissue homogeneity. 2. Insulation and precipitationprotein:
take test tube 3, number, press the table operation:
tube Reagent td width | < "140" >
C
liver tissue homogeneity
2.0 ml
2.0 ml
pre-boiled
liver tissue
2.0 ml
-
-
pH 7.6
phosphate Buffer
4.0 ml
4.0 ml
4.0 ml
orthodic acid
2.0 ml
2.0 ml
2.0 ml
-
4.0 ml
-
pre-boiled
muscle tissue Slurry
4.0 ml
-
4.0 ml
43 degrees C water bath insulation for 60 minutes
15% triclosan acetate | 3.0 ml | < After >
ef
and "> filtering in three test tubes, respectively.< td width"94" >
1
2
3
td width
5.0 ml
5.0 ml
5.0 ml
0.1mol/L I2-KI
3.0 ml
3.0 ml
3.0 ml
10% NaOH
3.0 ml
3.0 ml
3.0 ml
the amount of ketone produced by the liver (mmol/g) - (C-A) ×na2S2O3 moles× 1/6
the amount of ketone used by the muscles (mmol/g) - (C-B) × the number of moles in Na2S2O3 ×1/6
A: The number of Na2S2O3ml consumed by titring sample 1.
B: Titring sample 2 consumes Na2S2O3ml.
C: The number of Na2S2O3ml consumed by titring sample 3."thinking"
Why can ketones be oxidized and used only in extra-liver tissue
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