practical biochemistry 2

• quantitative testing for reducing sugar

  • the amount of precipitate will increase
  • amount of copper ions remaining in solution will decrease

  using colorimetry we can quantify how concentration of sugar in original sample by assessing the variables change
• colorimeter

  works by shining light through sample

  • we would use a centrifuge to separate the precipitate and any excess Benedict's solution
  • using a pipette we can take the supernatant and place in a cuvette, which is then placed in a colorimeter

  leaving greasy fingerprint can affect the transmission of light
• colorimetry 2

  for greater accuracy we can use red colour filter

  • red filter can detect percentage transmission + solution reflects blue light but absorbs red light
  • if there's a lot of unreacted copper sulphate, supernatant is still blue, absorption of red light is high and percentage transmission is low
  • if there's little of unreacted copper sulphate, supernatant is less blue, absorption of red light is low and percentage transmission is high
• the device is usually zeroed between each reading by placing a 'blank' sample to reset absorption/transmission to 100%
• creating a calibration curve

  colorimeter gives us a semi-quantitative test for sugar, as we can compare amount of sugar in different samples

  to find exact amounts we need a calibration curve

  1. first, take a series of known concentrations of reducing sugar
  2. using a sample of each, carry out Benedict's solution
  3. use a colorimeter to record percentage transmission of light through each supernatant
  4. plot a graph to show 'transmission of light' against the concentration of reducing sugar
• providing a calibration curve, you can use with other 'unknown' samples to determine concentration of sugar in original
• biosensors - convert biological or chemical variable which cannot be easily measure into an electrical signal

  e.g colorimeter