Quantative analysis of caffeine content

Caffeine was extracted from decaffeinated java and normal tea utilizing DCM as the dissolver and compared with prepared solutions of caffeine utilizing rearward stage HPLC. The by experimentation determined consequence was a caffeine concentration of 7.598? g/ml for decaffeinated java compared with 5.9? g/ml mention and 0.4049mg/ml for tea against 0.26mg/ml [ 1 ] * .


Caffeine ( 1 ) is the most normally consumed non-prescription stimulation. It is a member of the xanthine ( 2 ) household [ 1 ] and is structurally related to other works alkaloids such as theobromine ( 3 ) , normally found in chocolate beans and cocoa.

The mean individual in the UK drinks over thee cups of tea a twenty-four hours, [ 2 ] and so quantatively cognizing the sum of caffeine that is contained in a cup of tea is peculiarly of import. Caffeine Acts of the Apostless on certain adenosine receptors in the encephalon [ 3 ] , although there is grounds that action on these specific receptors may excite others such as Dopastat receptors, normally associated with wages [ 4 ] . More chronic effects of caffeine ingestion include crossness, impaired concentration, muscle-aches, sickness, purging and blurred vision [ 5 ] .

HPLC denotes high force per unit area liquid chromatography. In this experiment, change by reversal stage HPLC was used, intending that partly polar organic dissolvers were used as a nomadic stage against a non-polar stationary stage, in this instance, octadecyl silane ironss [ 6 ] . HPLC is a utile tool in analyzing rapidly and accurately comparative copiousnesss of assorted chemicals contained within a sample.

The experiment was undertaken to measure the caffeine content in an mean cup of tea and an mean cup of decaffeinated java.


The experiment was carried out harmonizing to the University of Bristol Second Year Laboratory Manual. Consequences were analysed by contrary HPLC utilizing an isocratic dissolver system dwelling of 10 millimeters ammonium ethanoate, methyl alcohol and tetrahydrofuran at a ratio of 90:8.5:1.5. The non-polar stage featured octadecyl silane ironss at a atom size of 5? m. The injection volume was 10? cubic decimeter, flow rate was 1.5ml min-1 with UV monitoring at 273nm.

Preparation of Tea

One tea bag was added to 200ml of boiling deionised H2O and boiled ( ca. 10 proceedingss ) with occasional agitation. Tea bag was removed and solution allowed to chill to room temperature. Sodium chloride ( 5g, 0.856moles ) added with Ca hydrated oxide ( 1g, 0.0135moles ) twice filtered by B & A ; uuml ; chner filtration. 20ml transferred to extraction funnel, extracted with DCM ( 20ml ) three times. Magnesium sulfate added to dry DCM thoroughly of all H2O. Evaporated to dryness via rotary vaporization. Staying solid dissolved in DCM and made up to 20ml.

Preparation of Decaffeinated Coffee

1.50g of Decaffeinated Coffee granules was added to 200ml of boiling H2O and allowed to chill to room temperature. Calcium hydrated oxide added ( 1g, 0.0135moles ) to decaffeinated java solution. Solution twice filtered via B & A ; uuml ; chner filtration. 20ml of solution transferred to extraction funnel and extracted with three equivalents of DCM ( 20ml ) . Magnesium sulfate added to dry the DCM, B & A ; uuml ; chner filtered and evaporated to dryness via rotary vaporization. Staying solid dissolved in DCM and made up to 20ml.

Consequences and Discussion

The tea solution was originally brown and crystalline, and the java was black. Addition of Ca hydrated oxide changed the coloring material of both solutions to an opaque dark brown. Tannins are precipitated by lime H2O, and so this is likely to be what precipitated out in the tea and java solution. [ 7 ] This precipitate was so removed by Buchner filtration. It was besides found that both solutions, java particularly, readily formed emulsions and bubbles in the extraction funnel, and so a big measure of Mg sulfate was needed to take all H2O from the extractions. This may hold interfered with the full extraction of the caffeine.

The soaking up strength for Tea was 7108493 units. Substituting this into equation 1 allows us to cipher the caffeine concentration in Tea. The solved concentration is 0.4049mg/ml.

The soaking up strength for Decaffeinated Coffee was 79233 units. Substituting into equation 1 allows us to cipher the caffeine concentration in decaffeinated java. The solved concentration is 0.007598mg/ml.

Mistakes in solution concentration

The mistake for the balance used to mensurate out the caffeine pulverization for the standard solutions was ±0.00005g. The mistake for the 20ml volumetric flasks used was ±0.1ml. Substituting these to happen maximal and minimal values into the moles equation allows for accurate mistake analysis.

Substituting these values into the informations and plotting onto a graph grants the ability to pull the same graph above with mistake boxes. It besides allows for upper limit and minimal gradients to be drawn with the upper limit and minimal values. From this information precise mistakes in the concentration of tea and decaffeinated java can be calculated.

Substituting maximal values into a graph enables the gradient and stop to be calculated, and therefore the positive mistake in caffeine concentration of the tea and decaffeinated java solutions. The same can be done with minimal values.

Substituting in values of optical density for tea gives a maximal value of 0.4069mg/ml and minimal value of 0.4029mg/ml. For decaffeinated java the maximal value of 7.639? g/ml and minimal value of 7.558? g/ml.

Literature values for the caffeine content of decaffeinated java are in the part of 0.5-1.5mg per 177.44ml1, which is between 0.0003-0.009mg/ml. This is near to the recorded value of 7.598? g/ml ± . The literature values for the caffeine content in tea are 15-75mg/ml per 177.44ml, which is between 0.088-0.440mg/ml. This is near to the recorded value of 0.4049mg/ml ± . Both of the obtained values are within the scope given in mention ; nevertheless both consequences are in the upper terminal of values that should be obtained. This could be due to hapless measuring of either the mention caffeine extremums or instant java measuring.


The isolation of caffeine from tea and instant java was successful and the values obtained from the quantative analysis of a sample by contrary HPLC compared favorably to cite values. More accurate values for caffeine content could be obtained by larger measures of instant java samples and by repeat of checks affecting caffeine extraction from tea. The importance of specifying the measure of caffeine in decaffeinated java is of import since the concentration must stay low.


I would wish to thank my demonstrators Jack Davidson and Chun Zhu for assisting me with the experiment and my lab spouse Ben Gough.


  1. B. A. Weinberg, B. K. Bealer, The universe of caffeine: the scientific discipline and civilization of the universe ‘s most popular drug, Routledge ( 2001 ) pp. 216.
  2. G. A. Spiller, Caffeine, CRC Press, ( 1998 ) pp. 38.
  3. U. Gupta, Caffeine and behaviour: current positions and research tendencies, CRC Press ( 1999 ) pp. 1.
  4. R. S. J. Frackowiak, Human encephalon map, Academic Press ( 2004 ) pp. 446.
  5. B. A. Weinberg, B. K. Bealer, The universe of caffeine: the scientific discipline and civilization of the universe ‘s most popular drug, Routledge ( 2001 ) pp. 304.
  6. University of Bristol, School of Chemistry, Level 2 Teaching Laboratory Manual 2009/2010.
  7. P. J. Van Soest, Nutritional ecology of the ruminant, Cornell University Press ( 1994 ) pp. 204-205.

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