title: Determination of caffeine, theobromine, and theophylline in standard reference material 2384, baking chocolate, using reversed-phase liquid chromatography.
abstract: A rapid and selective isocratic reversed-phase liquid chromatographic method has been developed at the National Institute of Standards and Technology to simultaneously measure caffeine, theobromine, and theophylline in a food-matrix standard reference material (SRM) 2384, Baking Chocolate. The method uses isocratic elution with a mobile phase composition (volume fractions) of 10% acetronitrile/90% water (pH adjusted to 2.5 using acetic acid) at a flow rate of 1.5 mL/min with ultraviolet absorbance detection (274 nm). Total elution time for these analytes is less than 15 min. Concentration levels of caffeine, theobromine, and theophylline were measured in single 1-g samples taken from each of eight bars of chocolate over an eight-day period. Samples were defatted with hexane, and beta-hydroxyethyltheophylline was added as the internal standard. The repeatability for the caffeine, theobromine, and theophylline measurements was 5.1, 2.3, and 1.9%, respectively. The limit of quantitation for all analytes was <100 ng/mL. The measurements from this method were used in the value-assignment of caffeine, theobromine, and theophylline in SRM 2384.
authors.affiliation: United States Department of Commerce, National Institute of Standards and Technology, Chemical Science and Technology Laboratory, Gaithersburg, Maryland 20899-0001, USA. jbthomas@nist.gov
authors.forename: Jeanice Brown
authors.initials: JB
authors.lastname: Thomas
authors.forename: James H
authors.initials: JH
authors.lastname: Yen
authors.forename: Michele M
authors.initials: MM
authors.lastname: Schantz
authors.forename: Barbara J
authors.initials: BJ
authors.lastname: Porter
authors.forename: Katherine E
authors.initials: KE
authors.lastname: Sharpless
mesh_terms: D002099:Cacao / Q000737:chemistry*; D002110:Caffeine / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D012015:Reference Standards; D013805:Theobromine / Q000032:analysis*; D013806:Theophylline / Q000032:analysis*
chemical_list: D002110:Caffeine; D013806:Theophylline; D013805:Theobromine