Determination of Caffeine in Different Types of Coffee and Tea by High Performance Liquid Chromatography (HPLC)

Chen, Chee Shien (2022) Determination of Caffeine in Different Types of Coffee and Tea by High Performance Liquid Chromatography (HPLC). Final Year Project (Bachelor), Tunku Abdul Rahman University College.

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Abstract

A reversed-phase high performance liquid chromatography (RP-HPLC) method was developed and validated for the quantitative analysis of caffeine content in 40 different types of tea and coffee samples. Ethanol extracted caffeine was separated through C18 reversed phase column by using 0.10 % formic acid solution and acetonitrile with ratio of 80: 20 as mobile phase at flow rate of 1.0 mL/min. The caffeine peak would appear at the retention time of 4.0 ± 0.1 minutes when the diode-array detection (DAD) detector was set at a wavelength of 278 nm. The method developed was validated through 6 parameters involving specificity, accuracy, precision, linearity, limit of detection (LOD), and limit of quantitation (LOQ). The percent recoveries obtained were within the acceptable range of 80.00 % to 120.00 %. The relative standard deviations (% RSD) for intra-day and inter-day precisions were less than 2.00 %. A positive correlation between the peak areas and the caffeine concentrations was observed as the correlation coefficient (R2) of 0.9995 was obtained. The LOD obtained was 0.05 μg/mL and the LOQ obtained was 0.16 μg/mL. After validation, the HPLC method developed was used to determine the concentration of caffeine in different types of tea and coffee samples. For the tea samples, the caffeine content in the tea beverages ranged from 0.00010 % w/v to 0.00023 % w/v, whereas the caffeine content in the tea leaves ranged from 0.13 % w/w to 0.82 % w/w. In contrast, for the coffee samples, the caffeine content in the coffee beverages ranged from 0.00028 % w/v to 0.00072 % w/v, whereas instant coffee had a caffeine content ranging from 0.08 % w/w to 0.96 % w/w. Besides, the concentration of caffeine in the coffee beverages was much higher than that of tea beverages, whereas the caffeine content in instant coffee was lower than that of the tea leaves. Furthermore, the caffeine content in the solid samples was found to be greater than that of the liquid samples for both tea and coffee. In short, high consumption of caffeine is harmful to human health, and hence, the amount of caffeine in caffeinated products must be regulated strictly