Analytical Chemistry, 2016, 88: 11222-11228.

Wiring Bacterial Electron Flow for Sensitive Whole-Cell Amperometric Detection of Riboflavin

Rong-Wei Si1,2, Yuan Yang1,2, Yang-Yang Yu1,2, Song Han2, Chun-Lian Zhang1,2, De-Zhen Sun1,2, Dan-Dan Zhai1,2, Xiang Liu1,2, Yang-Chun Yong1,2

1Biofuels Institute and 2School of the Environment, Jiangsu University, 301 Xuefu Road, Zhenjiang 212013, Jiangsu Province, China

 

Abstract

A whole-cell bioelectrochemical biosensing system for amperometric detection of riboflavin was developed. A “bioelectrochemical wire” (BW) consists of riboflavin and cytochrome C between S. oneidensis MR-1 and electrode was characterized. Typically, strong electrochemical response was observed when riboflavin was added to reinforce this BW. Impressively, the electrochemical response of riboflavin with this BW was over 200 times higher than that without bacteria. Uniquely, this electron rewiring process enabled the development of a biosensing system for amperometric detection of riboflavin. Remarkably, this amperometric method showed high sensitivity (LOD=2.2 nM, S/N=3), wide linear range (5 nM~10 μM, three orders of magnitude), good selectivity and high resistance to interferences. Additionally, the developed amperometric method featured good stability and reusability, and was applied for accurate and reliable determination of riboflavin in real conditions including food, pharmaceutical, and clinical samples without pretreatment. Both the cost-effectiveness and robustness make this whole-cell amperometric system ideal for practical applications. This work demonstrated the power of bioelectrochemical signal amplification with exoelectrogen and also provided a new idea for development of versatile whole-cell amperometric biosensors.

Key words: Bioelectrochemical systemï¼›Biosensor; Riboflavin; Extracellular electron transfer; Shewanella 

PMID: 27750415

 

Supplement:

Riboflavin is an important and essential nutrient to human as it played crucial roles in different biological processes and it could not be synthesized by human body. So, humanbeing is solely dependent on riboflavin uptaking from food or medicine. Moreover, both of deficiency and excess of riboflavin in human body would cause a series of diseases even cancer. Therefore, riboflavin was considered as a key clinical biomarker. Current methods for riboflavin analysis include HPLC, capillary electrophoresis, and conventional voltammetry. However, efficient diagnosis is still suffered from the problems associated with these methods including expensive equipment requirement, complicated procedure, low sensitivity, etc.

 

Our lab developed a new analytical tool for sensitive detection of riboflavin in clinical samples. This new biosensor is consists of a conventional three-electrode electrochemical cell system as the signal collection system, Shewanella cells as the recognition module and signal generation module, the signal collected can be directly recorded by a simple potentiostat (Fig. 1). It was found that the surface-bounded electroactive protein cytochrome C of Shewanella could selectively recognize the riboflavin molecule, and activate the inwards electron transfer flow from the electrode to the cells, which was also driven by the intracellular fumarate reduction. With the saturated fumarate, the intensity of the inwards electron flow (under an amperometric condition) was dependent on the concentration of riboflavin in the system. Therefore, riboflavin could be simply detected amperometrically by this whole-cell biosensing system.

 

 

Fig.1. Schematic of the developed biosensing system. EAB indicates the electroactive bacteria (here is S. oneidensis MR-1). CE, counter electrode; RE, reference electrode; WE, working electrode.

 

More impressively, the limit of detection (LOD) of this biosensing system estimated is ~2.2 nM, which is much lower than previous reported electrochemical methods for riboflavin detection. The linear detection range obtained by the newly developed biosensor is ranging from 5 nM to 10 μM, which is also superior to other methods and useful for practical application. Furthermore, this kind of biosensor is robust enough for clinical diagnosis as it showed excellent storage stability, specificity and resuablity. Uniquely, this biosensor showed high resistance to bacterial contamination which is usually impossible for other whole-cell biosensor, because. most of the bacteria in the environment can not generate electric response under the biosensing condition. This biosensing system featured as simple equipment requirement (simplest potentiostat), simple procedure (whithot complicated pretreatment step), high sensitivity and robustness, which would be promising for practical application. The idea of wiring electrode with whole-cell for bioelectrochemical sensing system development opened up new possibility for biosensor design and would show great impact in the field of whole-cell biosensor.

 

Acknowledgements: This work was partially supported by the National Natural Science Foundation of China (NSFC 51578266), Natural Science Foundation of Jiangsu Province for Distinguished Young Scholars (BK20160015).

 

Contact:

Yang-Chun Yong, Ph.D.

Professor

School of Environment and Safety Engineering,

Jiangsu University

301 Xuefu Road, Zhenjiang 212013, Jiangsu Province, China

Email: ycyong@ujs.edu.cn