Date of Award


Degree Name

MS in Agriculture - Dairy Products Technology


Dairy Science


Rafael Jimenez-Flores


Coffee is a popular and well-loved beverage consumed worldwide by millions of people every day. While most patrons of coffee do so because of its unique and satisfying taste, consumers may be unaware of the potential beneficial health effects it also imparts. The antioxidants found in green coffee beans collectively known as chlorogenic acids (CGA) and caffeine are two of the most abundant bioactive compounds present in coffee. Both these bioactive compounds have been implicated in many studies to impart a wide range of health benefits, from reducing the risk of Type 2 diabetes, to their use as aides in weight management. Indeed, epidemiological studies on people who consume moderate amounts of coffee on a regular basis have unanimously shown benefits to overall health.

While caffeine and CGA are naturally occurring compounds in coffee, their potential in conferring beneficial health effects warrant research into other potential food matrixes that can be used to bind and deliver these bioactive compounds into foods that do not naturally contain them. Milk proteins, specifically caseins, have been shown to be excellent vehicles to both bind and deliver sensitive bioactive compounds of various chemical and physical properties. Caseins have been shown in numerous studies to successfully bind to molecules such as vitamin D2, ω-3 polyunsaturated fatty acids (DHA), and iron to name a few. Because caseins exhibit high versatility in binding a variety of molecules, caseins were the milk protein of choice for the experiments in this thesis.

Polyphenols have been the subject of many studies on its binding capacity with milk proteins, but research on the binding capacity of caffeine with caseins is limited. Therefore, the objectives of this thesis are threefold: 1) develop, optimize and validate an HPLC method for the accurate and simultaneous determination of caffeine and CGA, 2) establish a procedure by which caffeine and CGA bind to sodium caseinate, and 3) determine the optimal treatment conditions of pH and temperature to increase binding interactions and speculate on the mechanism of binding for each bioactive compound.

A reversed phase HPLC (RP-HPLC) method was developed and subjected to validation studies with good results in linearity (caffeine R2 = 0.9992, CGA R2 = 0.9995) and precision (RSD of caffeine <1%, RSD of CGA <2%). The developed method also demonstrated selectivity for caffeine and CGA. This method was then used to analyze sodium caseinate samples containing caffeine and 5-CGA. The results from these studies have shown that binding interactions between caffeine and sodium caseinate are temperature dependent (p < 0.01) whereas binding interactions between CGA and sodium caseinate are influenced by both pH and temperature (p < 0.01). Elucidating the binding mechanisms of caffeine and CGA to sodium caseinate and providing a sensitive analytical technique by which these compounds can be accurately quantified may facilitate future research involving the use of caffeine and CGA in many other facets, as well as promoting its increased use in the dairy industry.