Hill reaction (Hill reaction) is a reaction in which isolated chloroplasts of green plants decompose water in the presence of light, releasing oxygen and at the same time reducing the electron acceptor. The oxidant 2,6-dichlorophenol indophenol is a blue dye that accepts electrons and H+ and is reduced to colorless, which can be observed directly as a color change, or spectrophotometrically, and the amount of reduction can be measured accurately.

Operation method

Observational experiments on Hill's reaction

Principle

Hill reaction (Hill reaction) is a reaction in which isolated chloroplasts of green plants decompose water in the presence of light, releasing oxygen and at the same time reducing the electron acceptor. The oxidant 2,6-dichlorophenol indophenol is a blue dye that accepts electrons and H+ and is reduced to colorless, which can be observed directly as a color change, or spectrophotometrically, and the amount of reduction can be measured accurately.

Materials and Instruments

Spinach
2-6-Dichlorophenol Indophenol Quartz sand Trimethylolaminomethane Sucrose NaCl
Mortar and pestle Graduated test tube Funnel Gauze Small beaker Scissors

Move

I. Materials, instrumentation and reagents

1.Materials

Fresh leaves of spinach or other green plants.

2.Apparatus

Mortar, graduated test tube, funnel, gauze, small beaker, scissors.

3.Reagents

(1) 0.1% 2,6-dichlorophenol indophenol, quartz sand.

(2) Extraction buffer: containing 0.05 M tris (trihydroxymethylaminomethane), pH 7.5, 0.4 M sucrose, 0.01 M NaCl;

II. Experimental steps

1. First take ice into a stainless steel basin and put it into a plastic bag, then place the pre-cooled mortar on the ice.

2. Take 12 grams of fresh spinach leaves, wash and dry them, remove the midrib and cut them into pieces and put them into the mortar on ice, add 10 ml of (pre-frozen) extract.

3. Grind and crush the cells quickly (make sure to crush thoroughly), then mix with about 10 ml of the frozen extract to obtain a suspension. Pass the suspension through three layers of gauze and filter into a small beaker.

4. Load the filtrate into a centrifuge tube and centrifuge at 1000 rpm for 3-5 minutes (the tube must be leveled), load the supernatant solution into another centrifuge tube (try to remove the impurities, preferring to lose part of the solution rather than transferring the precipitate into the centrifuge tube), and discard the precipitate (unbroken cells and tissue residue). The chloroplast suspension is obtained.

5. Take two test tubes and add 5 ml of chloroplast suspension to each tube.

6. Add 5-6 drops of 0.1% 2,6-dichlorophenol indophenol solution and shake well. Place one of the test tubes in direct light and the other in the dark and note the change in color of the solution in the sunlit tube. 5-8 M in later, remove the dark tube and compare the change in color of the solution in the two test tubes and explain why.

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