Molecular Specific and Sensitive Detection of Pyrazinamide and Its Metabolite Pyrazinoic Acid by Means of Surface Enhanced Raman Spectroscopy Employing In Situ Prepared Colloids

Anna Muehlig,Izabella J. Jahn,Patricia Sheen,Karina Weber,Jan Heidler,Martin Jahn,Dana Cialla-May,Mirko Zimic,Juergen Popp

doi:10.3390/app9122511

Abstract

The prodrug pyrazinamide (PZA) is metabolized by the mycobacteria to pyrazinoic acid (POA), which is expelled into the extracellular environment. PZA resistance is highly associated to a lack of POA efflux. Thus, by detecting a reduction of the concentration of POA in the extracellular environment, by means of lab-on-a-chip (LoC)-SERS (surface-enhanced Raman spectroscopy), an alternative approach for the discrimination of PZA resistant mycobacteria is introduced. A droplet-based microfluidic SERS device has been employed to illustrate the potential of the LoC-SERS method for the discrimination of PZA resistant mycobacteria. The two analytes were detected discretely in aqueous solution with a limit of detection of 27 µm for PZA and 21 µm for POA. The simultaneous detection of PZA and POA in aqueous mixtures could be realized within a concentration range from 20 μm to 50 μm for PZA and from 50 μm to 80 μm for POA.

Highlights

Tuberculosis (TB) is a disease affecting 10.0 million people globally each year [1], with drug resistance becoming a growing threat
Pyrazinamide (PZA, 97.5%), pyrazinoic acid (POA, 99%), AgNO3 (99.9999% trace metals basis), hydroxylamine hydrochloride (99.999% trace metals basis), hydrazine monohydrate (64–65%, reagent grade, 98%), sodium citrate (ACS reagent, ≥99.0%) and chile salpeter (NaNO3, ≥99.0%) were purchased from Sigma Aldrich, Darmstadt, Germany and used as received
The potential of LoC-SERS as a spectroscopic based method was the SERS spectra of PZA and POA were determined, e.g., for PZA specific marker bands are centered demonstrated for the simultaneous detection of PZA and POA in a microfluidic device

Summary

Introduction

Tuberculosis (TB) is a disease affecting 10.0 million people globally each year [1], with drug resistance becoming a growing threat. Sci. 2019, 9, 2511 worldwide, exacerbated by HIV-TB co-infection [2] and the emergence of multidrug-resistant (MDR) and extensively drug resistant (XDR) TB strains in both industrialized and developing countries [2,3]. TB is a curable disease; the mortality rate associated with this medical condition has considerably increased during the last years. The newly emerging MDR mycobacteria strains lead to a doubling of the MDR TB cases that are hard to treat. The early and correct identification of the drug susceptibility of the TB strain affecting a patient is crucial for a positive anti-tuberculous treatment outcome

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Conclusion