Experiment To Establish The Link Between Light Dependent And Independent Reactions

520 words - 2 pages


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Practical Laboratory Experiment John SimmonsAn experiment to demonstrate the link between light dependent and independent reactions in plant leaf matter.IntroductionThis practical experiment is to determine whether leaf material may be used to show photosynthesis.In light dependent reactions, NADP is the reducing agent (it loses, or donates an electron and is oxidized) which is then used to produce reducing power for light independent reaction.Isolating chloroplasts, the cell organelle which has the primary function is photosynthesis, will test this hypothesis, and subjecting it to lit and dark environments in order to record and analyse the results.Solutions of other materials used in the experiment will also be tested as a control.MethodThe chloroplasts is isolated by grinding the leaf material (spinach),using a pestle and mortar, and the resulting pulp or brei, is sieved and equally divided into centrifuge tubes and separated by centrifugal force for ten minutes.Next the supernatant, the residual liquid above the chloroplast pellet, is poured into a separate container and the chloroplast is suspended with isolation medium and remixed. This is then kept cold until use.Five labelled tubes are then prepared using the suspended leaf material, numbered one to five.DCPIP (2,6, Dichlorophenolindophenol), is a blue dye, acts as an electron acceptor and becomes colourless when reduced. Therefore, when this is added to isolated chloroplasts, any reducing agent produced by them may be detected.The DCPIP should be used at room temperature.Two of the tubes containing leaf material are placed under light and one in darkness. Readings are then recorded using a colorimeter, noting time and environment. Other tubes containing no chloroplast pellets are used as a control.The table below (a) records the contents and environment of the tubes used in the experiment.

Leaf extract/cm3


Isolation medium

Distilled water


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