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.