Simultaneous estimation of xanthine alkaloids (Theophylline, Theobromine and Caffeine) by High-Performance Liquid Chromatography
Methylxanthines are mainly a group of phytochemicals which are derived from purine base xanthine. These xanthines are obtained from plant as a result of secondary metabolism. There are various physiological actions that have been attributed to these derivatives in neurogenerative diseases, respiratory diseases, cancer and diabetes. The aim of this study is to develop a suitable qualitative and quantitative method for these xanthine derivatives. HPLC method is suitable for simultaneous estimation of methylxanthines, based on their physicochemical properties. Theobromine, Theophylline and Caffeine that belongs to alkaloids possess their economic effects. They have various stimulant effects on cardiovascular system, gastrointestinal system, respiratory system, central nervous system etc. that results in increased motivation to work, increased energy and increased alertness. High performance liquid chromatography is used for simultaneous determination of theophylline, theobromine and caffeine from different tea leaves.
2. Venkatesh S, Swamy MM, Reddy YS, Suresh B, Sethuraman M. A simple method for determination of caffeine content in tea samples. Anc Sci Life [Internet]. 1994;14(12): 358. Available from:
3. Sai V, Chaturvedula P, Prakash I. The aroma , taste, color and bioactive constituents of tea. J Med Plants Res. 2011;5(11):2110-24.
4. Sharma V, Gulati A, Ravindranath SD, Kumar V. A simple and convenient method for analysis of tea biochemicals by reverse phase HPLC. J Food Compos Anal. 2005;18(6):583-94.
5. Theobromine [Internet]. Sciencedirect; 2010 [cited 2019 Feb 26]. Available from:
6. Raghavendra HL, Kumar SVP, Kekuda TRP, Ejeta E, Molla B, Anilakumar KR, et al. HPLC Method for Chemical Composition and In vitro Antioxidant Activity of Camellia sinensis Linn. .Anal Chem Lett. 2013;1(5-6):361-9.
7. Tea varieties [Internet]. Brand India; 2019 [cited 2019 Feb 25]. Available from:
8. Kokate CK. Purohit P, Gokhale SB. A Text Book of Pharmacognosy. 2nd ed. New Delhi; Nirali Publication; 1990 .p.3.72.
9. Erickson J. Determination of the concentration of caffeine, theobromine, and gallic acid in commercial tea samples. Concordia Coll J Anal Chem [Internet]. 2011 [cited 2019 Feb 22]; 2:315. Available from:
10. Bispo MS, Veloso MCC, Pinheiro HLC, De Oliveira RFS, Reis JON, Andrade JBD. Simultaneous determination of caffeine, theobromine, and theophylline by high-performance liquid chromatography. J Chromatogr Sci. 2002; 40(1):45-8.
11. Bucklin MH, Groth CM. Theophylline. Encycl Toxicol 3rd ed. 2014; (May 2013):530-2.
12. Srdjenovic B, Djordjevic-Milic V, Grujic N, Injac R, Lepojevic Z. Simultaneous HPLC determination of caffeine, theobromine, and theophylline in food, drinks, and herbal products. J Chromatogr Sci. 2008; 46(2):144-9.
13. Roy A. A Review on the Alkaloids an Important Therapeutic Compound from Plants. Int J Plant Biotechnol. 2017; 3(2):1-9.
14. Rock L, Brunswick N. dear sophie youtube [Internet]. Google Search; 2005 [cited 2019 Feb 22]; 4290-302. Available from:
https://www.google.ca/webhp?sourceid=chrome-instant&ion=1&espv=2&es_th=1&ie=UTF-8#q=dear sophie youtube&es_th=1
15. Nadian NI, Uthumporn U. Determination of phenolic and antioxidant properties in Tea and spent Tea Under Various Extraction method and determination of Catechins, caffeine and Gallic Acid by HPLC. International journal on Advanced Science Engineering Information Technology. 2015; 5(1):158-164.
16. Adak M and Gabar MA. Green tea as a functional food for better health : A brief review. Research Journal of Pharmaceutical , Biological and Chemical Sciences. 2011; 2(2):645-64.
17. Komes D, Horžić D, Belščak A, Ganič KK, Baljak A. Determination of caffeine content in tea and maté tea by using different methods. Czech J Food Sci. 2009; 27(SPEC. ISS.):213-6.
18. Li HP, Tan HP, Yao S, Song H. Separation and preparation purine alkaloids from green tea waste. Asian J Chem. 2012; 24(12):5466-8.
19. Tamba A, Cioroiu B, Profire L, Lazar MI. HPLC Method for the Evaluation of Chromatographic Conditions for Separation of New Xanthine Derivatives. Cellulose Chemistry and Technology. 2014; 48:61-8.
20. Cabrera C, Giménez R, López MC. Determination of tea components with antioxidant activity. J Agric Food Chem. 2003; 51(15):4427-35.
21. Inti Ismail NN, Sapina Abdullah UU. Determination of Phenolic and Antioxidant Properties in Tea and Spent Tea Under Various Extraction Method and Determination of Catechins, Caffeine and Gallic Acid by HPLC. Int J Adv Sci Eng Inf Technol. 2016; 5(3):158.
22. De Sena AR, de Assis SA, Branco A. Analysis of theobromine and related compounds by reversed phase high-performance liquid chromatography with ultraviolet detection: An update (1992-2011). Food Technol Biotechnol. 2011; 49(4):413-23.
23. Horžić D, Komes D, Belščak A, Ganić KK, Iveković D, Karlović D. The composition of polyphenols and methylxanthines in teas and herbal infusions. Food Chem. 2009; 115(2):441-8.
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