SENSITIVE DETERMINATION OF 3,4-DIHYDROXY-L-PHENYLALANINE BY A CLOUD FUNNEL MUSHROOM (Clitocybe nebularis (Batsch), P. Kumm.) HOMOGENATE-BASED AMPEROMETRIC BIOSENSOR

Engin Asav

doi:10.23902/trkjnat.969982

Abstract

3,4-dihydroxy-L-phenylalanine (L-DOPA) is one of the precursor molecules for the biosynthesis of neurotransmitters in the brain. Monitoring of L-DOPA levels as a drug or biomolecule in biological fluids is crucial for the treatment of patients suffering from Parkinson’s Disease. This study aimed to construct a cloud funnel mushroom (Clitocybe nebularis (Batsch), P. Kumm.) tissue homogenate-based biosensor for precise and sensitive detection of L-DOPA in artificial plasma and urine. For this purpose, in the fabrication of the biosensor, tissue homogenate of C. nebularis was immobilized into a carbon paste electrode by using graphite, mineral oil, gelatine and glutaraldehyde. The amperometric signals corresponding to 600 s were recorded as response current for each L-DOPA concentration. All amperometric measurements were carried out at ⁻700 mV (versus Ag|AgCl). The present biosensor successfully detected L-DOPA with a linear dynamic range at 2.5-100 µM and Limit of Detection (LOD) value as 0.76 µM, as well as standard deviation as ±0.41 µM and coefficient of variation as 0.82% (n=16). Additionally, the determination of L-DOPA spiked in artificial plasma and urine was carried out successfully. The present work would be the first study that utilized C. nebularis tissue as a biosensor component.

Highlights

In total protein assay via the standard method of Bradford, the protein concentration of C. nebularis was determined as 0.421 mg/mL
The laccase activity determined via the ABTS method was calculated as 144.54 U/L according to the equation given above
The concentration of gelatin directly affected the signal rate, since gelatine acted as a slight barrier for oxygen and L-DOPA transport

Summary

Introduction

In the last two decades, the field of electrochemical biosensors has evolved rapidly by means of various types of transducers including amperometric (Ozcan & Aydin 2016), potentiometric (Rasmussen et al 2007), and voltammetric (Li et al 2015) along with bio-components such as tissues (Ozcan & Sagiroglu 2014), enzymes (Davletshina et al 2020), antibodies (Sayikli Şimşek et al 2015), microorganisms (Gao et al 2017) and DNA (Faria& Zucolotto 2019). Biosensor systems are widely used for the detection of several target molecules in the fields of medical diagnosis (Sun et al 2014), bioprocess control (Pontius et al 2020), In the construction of tissue-based electrochemical biosensors, tissue homogenates from various living organisms such as pigs (Thoppe Rajendran et al 2020), mushrooms (Sezgintürk & Dinçkaya 2012) and plant tissues including banana (Ozcan & Sagiroglu 2010), artichoke (Odaci et al 2004), Myrtle (Ayna & Akyilmaz 2018) are employed for detection of toxins (Sanders et al 2001), drugs (Thoppe Rajendran et al 2020), herbicides (Breton et al 2006) and phenolic compounds such as rutin (Zwirtes de Oliveira et al 2006), epinephrine (Felix et al 2006), caffeic acid (Fernandes et al 2007), catechol (Ozcan & Sagiroglu 2010) and dopamine (Ori et al 2014). C. nebularis was reported to have laccase gene (Luis et al 2004), neither determination of laccase activity nor utilization in biosensor construction have not been studied so far

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Conclusion