Potentiometric Detection of Serum Creatinine Utilizing Lead Dioxide and Carbon Nanotubes in a Single-Enzyme Reaction

Creatinine (CRE) serves as the end product of creatine, responsible for energy release in skeletal muscle. The baseline concentration of serum creatinine (SCr) typically ranges from 45 μM to 90 μM in females and 60 μM to 110 μM in males. Elevated SCr levels are commonly utilized in diagnosing acute kidney injury. Ongoing research focuses on developing reliable techniques for estimating SCr due to its critical role as a biomarker of significance. The Jaffe reaction, a colorimetric method utilizing picric acid, has traditionally been employed for clinical analysis of creatinine (CRE). However, this method presents several drawbacks, including the need for pH adjustment, interference issues, and, notably, the toxicity and explosiveness of picric acid. A prominent trend in CRE detection involves electrochemical enzyme sensors, offering enhanced sensitivity and selectivity. The inclusion of creatinine deiminase (CD) in the enzymatic approach addresses the aforementioned challenges, thereby improving sensor performance and selectivity. In this enzymatic method, CD reacts with CRE to produce NH₃, subsequently converted to NH₄⁺ and OH^- in an aqueous solution. The NH₄⁺ ion selective sensor and pH sensor then measure the concentration of CRE.<br/>Single-walled carbon nanotubes (SWCNTs) are widely acknowledged for their outstanding electrical properties, characterized by high carrier mobility and current-carrying capacity, coupled with impressive chemical, thermal, and mechanical attributes. Surface modification techniques can be applied to introduce diverse functional groups on the SWCNT surface, enhancing their suitability for various applications. The incorporation of metal oxide deposits can impart electrical, electrochemical, and biocompatible properties, making them valuable in sensor manufacturing. Among the diverse applications, electrochemical pH sensors stand out, and metal oxide modification presents several advantages. These include notable sensitivity, rapid response times, prolonged operational lifespan, minimal interference with other ions, cost-effectiveness, ease of maintenance, and adaptability for miniaturization in flexible systems. While various metal oxide pH sensors such as IrO₂, RuO₂, TiO₂, WO₃, and SnO₂ have been reported in recent years, their production often involves intricate and high-temperature processes. Lead dioxide (PbO₂) emerges as a noteworthy choice for pH sensors due to its excellent electrochemical properties and straightforward fabrication method. However, PbO₂-based pH sensors may exhibit deficiencies in oxygen or an excess of lead within the PbO₂, potentially impacting electrochemical reactions.<br/>In this investigation, we produced a thin film of lead dioxide deposited on carbon nanotubes (PbO₂/CNT) and developed a PbO₂/CNT ion-selective electrode (PbO₂/CNT/ISE). These sensors served as highly sensitive potentiometric biosensors for CRE, capitalizing on the one-step selective conversion of creatinine by creatinine deiminase. We conducted the detection of OH^- and NH₄⁺ in an aqueous solution, generated through an enzymatic reaction, utilizing PbO₂/CNT as a pH sensor and PbO₂/CNT/ISE as an NH₄⁺ selective electrode. The obtained results reveal a remarkable sensitivity of -75.56 mV log[CRE]^-1 and 64.62 mV log[CRE]^-1, with a calculated limit of detection of 0.06 μM and 0.13 μM, respectively, within the range of 10 to 400 μM. A selectivity test demonstrated robust discrimination against interfering materials in human serum, and the repeatability and long-term stability of our sensors were confirmed, indicating their high-level stability. Furthermore, recovery tests for CRE concentrations (10, 25, 50, 70 μM) in spiked human serum yielded results of 104%, 98%, 103%, and 96% for PbO₂/CNT, and 104%, 92%, 110%, and 97% for PbO₂/CNT/ISE. These findings underscore the reliability and reproducibility of our sensors in detecting CRE for clinical applications.