Determination of caffeine in beverages by high-performance liquid chromatography (HPLC

Determination of caffeine in beverages by high-performance liquid chromatography (HPLC)

 

Name :………………………………………………                                MSc Course :……………..

 

Date :………………………………

 

Marks Guideline

 

   
Complete the report as instructed.

Marks are given for:

·        Give a short introduction to HPLC and discuss the difference between accuracy and precision in analytical science

·        Plot graphs of the mean peak area and mean peak height for each of the standards against the concentration of the standard.  Graphs plotted correctly with appropriate title, units, equation, significant figures and line of best fit.

·        Use your graphs to determine the concentration of caffeine in the original beverage samples, use appropriate units and significant figures.  Show the calculations and include an error value.

·        Answering the questions on the proforma

·        Discuss your results, comment on the differences (if any) obtained using peak area and peak height and indicate which you think is the “better” result and why.

·        Indicate the potential sources of error and their magnitudes and give an overall estimate of the precision of the results obtained. Comment on how the method and results could be improved.

 
Total Possible Mark        20

 

 

 

 

 

SMA0020 – Determination of Caffeine in Beverages Using HPLC

 

 

 

 

 

The purpose of this experiment is to determine the concentration of caffeine contained in the beverage provided. A calibration curve will be constructed using caffeine standards, then the unknown sample will be measured and its caffeine content calculated.

Your graphs (drawn in Excel or a similar program) and your chromatograms must be attached to your report.

Part 1 – Calibration curve

Run each caffeine standard solution twice, starting with the lowest concentration. Record your measurements in the table below and plot a calibration graph for peak area vs. concentration. Copy and paste your graph in to this proforma.

 

Conc/ppm Peak area 1 Peak Area 2 Peak Height 1 Peak Height 2
10 93742 93013 9326 9326
25 222699 217540 21826 21826
50 490364 490344 49418 49418
75 726731 722776 72663 72663
100 903954 904227 90471 90471

 

Part 2 – Analysis of unknowns

Record the peak area and peak height for the unknown samples in triplicate.

 

Sample ID of unknown 1: Tea

 

Injection Peak area Peak Height
1 319403 30834
2 317675 30435
3 323347 31244
Average    
SD    

 

Calculate the concentration of caffeine in the unknown, including an error value:

 

Sample ID of unknown 2: Coffee

 

Injection Peak area Peak Height
1 437006 37455
2 414022 34879
3 373455 30318
Average    
SD    

 

Calculate the concentration of caffeine in the unknown, including an error value:

 

Sample ID of unknown 3: Red Bull

 

Injection Peak area Peak Height
1 644773 64823
2 666261 64345
3 655535 64155
Average    
SD    

 

Calculate the concentration of caffeine in the unknown, including an error value:

 

Questions

  1. Why were the calibration standards analysed in increasing order of concentration?
  2. Comment on the linearity of your calibration graphs
  3. Comment on the magnitude of the errors in your measurements of the unknown concentration.