Determination of Caffeine in Soda Introduction: High performance liquid chromatography (HPLC) is a type of chromatography which is very similar to liquid chromatography. In HPLC, however, the stationary phase column is more tightly packed than in other types of liquid chromatography. In this lab, the column is packed with C18 particles that are less than 10 m in diameter. The small diameter of the particles allows unprecedented resolution and high efficiency. Since the particles in the column are so small, it is necessary to pump the mobile phase through the column at a very high pressure. The pump keeps a precise flow rate so that the positions of the peaks in time can be used to identify the species in a sample. This is done by comparing the chromatographs of prepared standards of the particular species to be determined. The common peak is an indication of the standard. A small sample (100 Lx2) is injected into the injector port where the mobile phase moves it through the column. Each component, being different in physical composition, will move at a different rate through the C18 column. Thus, the components will be separated according to the size and shape of the molecules. The smallest and least hindered molecules will be eluted first since it is easiest for them to pass through the finely packed column. As each set of molecules elutes from the column, a detector (most often UV) recognizes it and records a peak. The area of this peak (in relation to the area of other peaks) is proportional to the concentration of that particular species in the sample. The identity can also be found by comparing the sample peaks to standards. Identical substances (peaks) will have identical retention times. Purpose: The purpose of this lab is to determine the amount of caffeine in a sample of soda. Materials: isocratic HPLC system vacuum 254 nm UV detector 60 mL syringe C18 column caffeine standards integrator 60:40 methanol : water (HPLC grade,degassed) 100 L syringes distilled water #42 filter paper HPLC grade methanol funnel soft drink sample computer printer Safety: Always wear safety glasses in the lab. Procedure: Preparation of soda samples 1. Obtain a soft drink sample.
Determination of Caffeine in Soda Introduction: High performance liquid chromatography (HPLC) is a type of chromatography which is very similar to liquid chromatography. In HPLC, however, the stationary phase column is more tightly packed than in other types of liquid chromatography. In this lab, the column is packed with C18 particles that are less than 10 m in diameter. The small diameter of the particles allows unprecedented resolution and high efficiency. Since the particles in the column are so small, it is necessary to pump the mobile phase through the column at a very high pressure. The pump keeps a precise flow rate so that the positions of the peaks in time can be used to identify the species in a sample. This is done by comparing the chromatographs of prepared standards of the particular species to be determined. The common peak is an indication of the standard. A small sample (100 Lx2) is injected into the injector port where the mobile phase moves it through the column. Each component, being different in physical composition, will move at a different rate through the C18 column. Thus, the components will be separated according to the size and shape of the molecules. The smallest and least hindered molecules will be eluted first since it is easiest for them to pass through the finely packed column. As each set of molecules elutes from the column, a detector (most often UV) recognizes it and records a peak. The area of this peak (in relation to the area of other peaks) is proportional to the concentration of that particular species in the sample. The identity can also be found by comparing the sample peaks to standards. Identical substances (peaks) will have identical retention times. Purpose: The purpose of this lab is to determine the amount of caffeine in a sample of soda. Materials: isocratic HPLC system vacuum 254 nm UV detector 60 mL syringe C18 column caffeine standards integrator 60:40 methanol : water (HPLC grade,degassed) 100 L syringes distilled water #42 filter paper HPLC grade methanol funnel soft drink sample computer printer Safety: Always wear safety glasses in the lab. Procedure: Preparation of soda samples 1. Obtain a soft drink sample.
HPLC_Determination%20of%20Caffeine%20in%20Soda.docx