Spectrophotometric determination of riboflavin concentration
Riboflavin, vitamin B2, is a water soluble vitamin. It needed in the co-enzyme FAD for instance. You can find riboflavin in liver, milk and egg. Riboflavin as a yellow green colour and can be determined by a spectrophotometer.
Task a. Determine a suitable wavelength for further measuring.
b. Make a calibration curve for riboflavin in the interval 5mg/cm3 – 20 mg/cm3.
c. Determine the content of riboflavin in a vitamin tablet.
Material Volumetric flask (1000 cm3 and 50 cm3), millimetre paper, spectrophotometer cells (quarts), spectrophotometer, 5 beakers (100 cm3)
Chemicals Riboflavin, 10% methanol solution, vitamin tablet
Procedure 1. Make a riboflavin standard solution for the whole group. 50.0 mg riboflavin is diluted to 1000 cm3 in 10% methanol solution. Calculate mg riboflavin per cm3.
2. Make from the standard solution three solutions of riboflavin in the interval 5mg/cm3 – 20 mg/cm3 with 10%methanol. Calculate the volume of standard solution you need to make 50 cm3 of the three solutions. Use the volumetric flask (50 cm3) and suitable pipettes. Mix well.
3. Look in the absorption spectrum of riboflavin below. Find the wavelength for the highest absorption and use that wavelength for further measurement.
4. Set the instrument at zero by using 10% methanol in both cells. Measure there after the absorbance for the three riboflavin solutions. Start with the least concentrated solution.
5. Write a calibration curve on a millimetre paper.
6. Pulverize a vitamin tablet in a mortar and take 15-20 mg (balance) to a 50cm3 volumetric flask. Add little 10% methanol and warm the flask in hot water so all are dissolved. Dilute with 10% methanol to 50 cm3. Mix well and measure the absorbance.
7. Calculate the mass of riboflavin in a tablet. Think of the dilutions. Compare with the value on the package.
Assessment DC DPP CE
Absorbansspectrum for Riboflavin: