Light Dependent Electron Transport, Using Dcpip

Light dependent electron transport, using DCPIP.

Introduction:

Photosynthesis is one of the most important processes in plants and therefore very important to us as humans, whether directly or indirectly. Photosynthesis employs structures called chloroplasts, which harness the sun’s light and through a series of reactions create sugars and useable energy such as ATP. There are two processes involved in photosynthesis the light dependent and the independent. This experiment explored the functions of the light dependent reactions in chloroplasts. The light dependent reaction and the chloroplasts harvest light energy to split H2O (absorbed by the plant) into protons, electrons and 02. Protons are used in a further reaction to create ATP, electrons are passed down the electron transport chain to yield NADPH and the O2 is expelled into the atmosphere. The aim of this experiment was to see/demonstrate how different conditions or treatments affect the light dependent electron transport of photosynthesis. This experiment used DCPIP in place of NADPH (the product normally produced in this reaction) as its characteristics/behavior match that of NADPH plus the added benefit of losing colour as it gains electrons. Therefore we can measure the rate of electron transport based on the changes in the colour of the solution. Of the six different treatments to be measured in this experiment only two of were expected to see a decrease in absorbance, the ‘Light’ and the ‘Red’, while the remainder of the tubes will have little to no change in absorbance.

Method:

Seven spectrophotometer tubes were numbered and solutions A-D were added according to the volumes shown in Table 1.  Tube 1 was capped and inverted several times. The spectrophotometer was calibrated using Tube 1, which contained chloroplasts and sucrose only, as the blank, to ensure that any changes in absorbance for the other treatments could be attributed to the reduction of the dye DCPIP.  At time zero (mins), absorbance was recorded for all treatments immediately after addition of DCPIP and mixing of contents.  Immediately following the time zero reading, tube 2 was wrapped in foil and tubes 6 and 7 were placed into larger tubes covered in red and green cellophane respectively.  Tubes 1-5 were also placed into larger tubes.  All tubes were then placed horizontally on ice, under lights.  At fifteen minute intervals, readings of absorbance were taken for all treatments, except for the dark tube which was kept wrapped in foil for 60 minutes, after which its absorbance was measured.”

Table 1.  Experimental design for the electron transport experiment.

Results:

There was an increase in absorbance where the reaction mixture was kept in the dark (Tube 2)(Fig.1) There was an overall decrease in absorbance where the reaction mixture was exposed to light for 60 minutes (Tube 3)(Fig.1). In Tube 4, where the reaction mixture contained boiled chloroplast suspension showed little to no decrease in absorbance overall (Fig.1) In the reaction mixture which had DCMU added (Tube 5) there was an increase in absorbance (Fig. 1). As for the coloured tubes, Red (Tube 6) and Green (Tube 7) showed small decreases in absorbance over the 60 minutes (Fig.1)