Simultaneous Quantification of Antioxidants Paraxanthine and Caffeine in Human Saliva by Electrochemical Sensing for CYP1A2 Phenotyping

Abstract

The enzyme CYP1A2 is responsible for the metabolism of numerous antioxidants in the body, including caffeine, which is transformed into paraxanthine, its main primary metabolite. Both molecules are known for their antioxidant and pro-oxidant characteristics, and the paraxanthine-to-caffeine molar ratio is a widely accepted metric for CYP1A2 phenotyping, to optimize dose–response effects in individual patients. We developed a simple, cheap and fast electrochemical based method for the simultaneous quantification of paraxanthine and caffeine in human saliva, by differential pulse voltammetry, using an anodically pretreated glassy carbon electrode. Cyclic voltammetry experiments revealed for the first time that the oxidation of paraxanthine is diffusion controlled with an irreversible peak at ca. +1.24 V (vs. Ag/AgCl) in a 0.1 M H2SO4 solution, and that the mechanism occurs via the transfer of two electrons and two protons. The simultaneous quantification of paraxanthine and caffeine was demonstrated in 0.1 M H2SO4 and spiked human saliva samples. In the latter case, limits of detection of 2.89 μM for paraxanthine and 5.80 μM for caffeine were obtained, respectively. The sensor is reliable, providing a relative standard deviation within 7% (n = 6). Potential applicability of the sensing platform was demonstrated by running a small scale trial on five healthy volunteers, with simultaneous quantification by differential pulse voltammetry (DPV) of paraxanthine and caffeine in saliva samples collected at 1, 3 and 6 h postdose administration. The results were validated by ultra-high pressure liquid chromatography and shown to have a high correlation factor (r = 0.994).

Recent Advances in Electrochemical Sensors for Caffeine Determination

“…Anastasiadi et al [ 53 ] used an anodically pretreated glassy carbon electrode as a sensor for the determination of caffeine and paraxanthine (PX) in human saliva. According to cyclic voltammetric measurements, the authors indicated that the shape and intensity of the current response depended on the pH value of the supporting electrolyte.…”

“…The peak potential of CF did not significantly change as the pH of the supporting solution increased, in contrast to other studies [ 83 ]. In addition, Anastasiadi et al [ 53 ] concluded that in a BR buffer solution at a higher pH value of 8, a larger background current occurs due to the discharge of the electrolyte. Guan et al [ 68 ] showed that the peak potential of CF remained unchanged with the increasing pH of PBS, indicating that protons were not included in the oxidation process.…”

Electrochemical sensors for analyte in saliva: recent update

“…For instance, electrochemical sensors have been developed for the detection of salivary pain killers, such as morphine (Eissa and Zourob, 2017), paracetamol (Campos et al , 2018), acetaminophen (Xie et al , 2021), tramadol (Arabali et al , 2020), aspirin (Diouf et al , 2020) and fentanyl (Mishra et al , 2022), which are widely used for pain management but also have a high potential for abuse and addiction. Similarly, electrochemical methods have been used for the a series of other pharmaceuticals, such as nicotine (Rajendran et al , 2022), diazepam (Hassan et al , 2020), cefuroxime (Kulapina et al , 2020), caffeine (Anastasiadi et al , 2020) and clonazepam (Ferreira et al , 2020).…”

Bioactive compounds in coffee and their role in lowering the risk of major public health consequences: A review

“…It is a plant alkaloid formed by methylation of the nitrogen atom of nicotinic acid by the catalytic effect of NaMN nucleotidase, NaR nucleotidase, and nicotinate methyl transferase (Arai et al, 2015;Mohamadi et al, 2017). It is found in the range of 0.6%-1.3% in F I G U R E 2 Chemical structure of caffeine and its metabolites (Anastasiadi et al, 2021).…”