Spectrophotometer

Objectives

1. To determine the wavelength of maximum absorption, Amax of Bromophenol blue.

2. To construct a standard concentration curve for Bromophenol blue.

3. To determine the concentration of the unknown Bromophenol blue solutions.

4. To determine the concentration of the different solutes, Bromophenol blue and Methyl orange, in a mixture.

Materials

Bromophenol blue, mixture of Bromophenol blue and Methyl orange solutions, micropipette, cuvette, test tubes, tips.

Methods

The experiments were carried out according to the procedures described in the practical manual.

Wavelength (nm) | Absorbance |

470 | 0.052 |

500 | 0.135 |

530 | 0.309 |

560 | 0.594 |

590 | 1.067 |

620 | 0.224 |

650 | 0.017 |

680 | 0.007 |

Table 1.2a: Bromophenol Blue Absorbance Value in Different Concentration at Wavelength of 590nm and 460nm.

Tube | 1 | 2 | 3 | 4 | 5 | 6 | A | B |

Distilled Water (ml) | 2.5 | 2.0 | 1.5 | 1.0 | 0.5 | 0 | - | - |

Bromophenol blue 10mg/L (ml) | 0 | 0.5 | 1.0 | 1.5 | 2.0 | 2.5 | - | - |

Concentration of Bromophenol blue (mg/L) | 0 | 2 | 4 | 6 | 8 | 10 | | |

Absorbance at 590nm (Amax of Bromophenol blue) | 0.000 | 0.220 | 0.430 | 0.645 | 0.850 | 1.053 | 0.331 | 0.189 |

Absorbance at 460nm | 0.000 | 0.017 | 0.023 | 0.036 | 0.048 | 0.065 | 0.002 | 0.005 |

Table 1.2b: Methyl Orange Absorbance Value in Different Concentration at Wavelength of 590nm and 460nm.

Tube | 1 | 2 | 3 | 4 | 5 | 6 |

Distilled Water (ml) | 2.5 | 2.0 | 1.5 | 1.0 | 0.5 | 0 |

Methyl orange 10mg/L (ml) | 0 | 0.5 | 1.0 | 1.5 | 2.0 | 2.5 |

Concentration of Methy orange (mg/L) | 0 | 2 | 4 | 6 | 8 | 10 |

Absorbance at 460nm | 0.000 | 0.157 | 0.307 | 0.461 | 0.611 | 0.764 |

Absorbance at 590nm (Amax of Bromophenol blue) | 0.000 | 0.001 | 0.002 | 0.003 | 0.003 | 0.005 |

Table 1.3

Tube | C |

Absorption of Mixture C at Amax of Bromophenol blue (BB), Ac at Amax BB | 0.532 mg/L |

Absorption of Mixture C at Amax of Methyl...