Selective Impedimetric Chemosensing of Carcinogenic Heterocyclic Aromatic Amine in Pork by dsDNA-Mimicking Molecularly Imprinted Polymer Film-Coated Electrodes.

Inspired by the easy intercalation of quinoxaline heterocyclic aromatic amines (HAAs) in double-stranded DNA (dsDNA), we synthesized a nucleobase-functionalized molecularly imprinted polymer (MIP) as the recognition unit of an impedimetric chemosensor for the selective determination of a 2-amino-3,7,8-trimethyl-3H-imidazo[4,5-f]quinoxaline (7,8-DiMeIQx) HAA. HAAs are generated in meat and fish processed at high temperatures. They are considered to be potent hazardous carcinogens. The MIP film was prepared by potentiodynamic electropolymerization of a pre-polymerization complex of two adenine- and one thymine-substituted bis(2,2'-bithien-5-yl)methane functional monomer molecules with one 7,8-DiMeIQx template molecule, in the presence of the 2,4,5,2',4',5'-hexa(thiophene-2-yl)-3,3'-bithiophene cross-linking monomer, in solution. The as-formed MIP chemosensor allowed for the selective impedimetric determination of 7,8-DiMeIQx in the 47 to 400 µM linear dynamic concentration range with a limit of detection of 15.5 µM. The chemosensor was successfully applied for 7,8-DiMeIQx determination in the pork meat extract as a proof of concept.

Oligonucleotide Analogs and Mimics for Sensing Macromolecular Biocompounds.

Living organisms create life-sustaining macromolecular biocompounds including biopolymers. Artificial polymers can selectively recognize biocompounds and are more resistant to harsh physical, chemical, and physiological conditions than biopolymers are. Due to recognition at a molecular level, molecularly imprinted polymers (MIPs) provide powerful tools to correlate structure with biological functionality and are often used to build next-generation chemosensors. We envision an increasing emergence of nucleic acid analogs (NAAs) or biorelevant monomers built into nature-mimicking polymers. For example, if nucleobases bearing monomers arranged by a complementary template are polymerized to form NAAs, the resulting MIPs will open up novel perspectives for synthesizing NAAs. Despite their usefulness, it is still challenging to use MIPs to devise adaptive biomaterials and to implement them in point-of-care testing.

Promoting bioanalytical concepts in genetics: A TATA box molecularly imprinted polymer as a small isolated fragment of the DNA damage repairing system.

We demonstrate that a new, stable, artificial TATA (T - thymine, A - adenine) box is recognized by amino acids recognizing the natural TATA box. Here, the former mimicked, as a minimal motif, oligodeoxyribonucleotide interactions with amino acids of proteins involved in repairing of damaged dsDNA. By electropolymerization, we molecularly imprinted non-labeled 5'-TATAAA-3' via Watson-Crick nucleobase pairing, thus synthesizing, in a one-step procedure, the hexakis[bis(2,2'-bithien-5-yl)] TTTATA and simultaneously hybridizing it with the 5'-TATAAA-3' template. That is, a stable dsDNA analog having a controlled sequence of nucleobases was formed in the molecularly imprinted polymer (MIP). The 5'-TATAAA-3' was by the X-ray photoelectron spectroscopy (XPS) depth profiling found to be homogeneously distributed both in the bulk of the MIP film and on its surface. The 5'-TATAAA-3' concentration in the 2.8(±0.2)-nm relative surface area, ~140-nm thick MIP film was 2.1 mM. The MIP served as a matrix of an artificial TATA box with the TATAAA-promoter sequence. We comprehensively characterized this artificial DNA hybrid by the polarization-modulation infrared reflection-absorption spectroscopy (PM-IRRAS) and X-ray photoelectron spectroscopy (XPS). Further, we examined interactions of DNA repairing TATA binding protein (TBP) amino acids with the artificial TATA box prepared. That is, molecules of l-phenylalanine aromatic amino acid were presumably engaged in stacking interactions with nucleobase steps of this artificial TATA box. The nitrogen-to­phosphorus atomic % ratio on the surface of the MIP-(5'-TATAAA-3') film increased by ~1.6 times after film immersing in the l-glutamic acid solution, as determined using the XPS depth profiling. Furthermore, l-lysine and l-serine preferentially interacted with the phosphate moiety of 5'-TATAAA-3'. We monitored amino acids interactions with the artificial TATA box using real-time piezoelectric microgravimetry at a quartz crystal microbalance (QCM) and surface plasmon resonance (SPR) spectroscopy under flow injection analysis (FIA) conditions.

Oligonucleotide Determination via Peptide Nucleic Acid Macromolecular Imprinting in an Electropolymerized CG-Rich Artificial Oligomer Analogue.

We devised and fabricated a chemosensor for determination of the genetically relevant 5'-GCGGCGGC-3' (G = guanine; C = cytosine) oligonucleotide. For that, we simultaneously electrosynthesized and electrode-immobilized a sequence-defined octakis(2,2'-bithien-5-yl) DNA hybridizing probe using both a "macromolecular imprinting in polymer strategy" and a sequence-programmable peptide nucleic acid (PNA) template. With electrochemical impedance spectroscopy (EIS) and surface plasmon resonance (SPR) transductions under stagnant-solution and flow injection analysis (FIA) conditions, respectively, we determined the above oligonucleotide with 200-pM EIS limit of detection. With its EIS-determined apparent imprinting factor of ∼4.0, the chemosensor was discriminative to both mismatched oligonucleotides and Dulbecco's modified Eagle's medium sample interferences.

Programmed Transfer of Sequence Information into a Molecularly Imprinted Polymer for Hexakis(2,2'-bithien-5-yl) DNA Analogue Formation toward Single-Nucleotide-Polymorphism Detection.

A new strategy of simple, inexpensive, rapid, and label-free single-nucleotide-polymorphism (SNP) detection using robust chemosensors with piezomicrogravimetric, surface plasmon resonance, or capacitive impedimetry (CI) signal transduction is reported. Using these chemosensors, selective detection of a genetically relevant oligonucleotide under FIA conditions within 2 min is accomplished. An invulnerable-to-nonspecific interaction molecularly imprinted polymer (MIP) with electrochemically synthesized probes of hexameric 2,2'-bithien-5-yl DNA analogues discriminating single purine-nucleobase mismatch at room temperature was used. With density functional theory modeling, the synthetic procedures developed, and isothermal titration calorimetry quantification, adenine (A)- or thymine (T)-substituted 2,2'-bithien-5-yl functional monomers capable of Watson-Crick nucleobase pairing with the TATAAA oligodeoxyribonucleotide template or its peptide nucleic acid (PNA) analogue were designed. Characterized by spectroscopic techniques, molecular cavities exposed the ordered nucleobases on the 2,2'-bithien-5-yl polymeric backbone of the TTTATA hexamer probe designed to hybridize the complementary TATAAA template. In that way, an artificial TATAAA-promoter sequence was formed in the MIP. The purine nucleobases of this sequence are known to be recognized by RNA polymerase to initiate the transcription in eukaryotes. The hexamer strongly hybridized TATAAA with the complex stability constant KsTTTATA-TATAAA = ka/kd ≈ 106 M-1, as high as that characteristic for longer-chain DNA-PNA hybrids. The CI chemosensor revealed a 5 nM limit of detection, quite appreciable as for the hexadeoxyribonucleotide. Molecular imprinting increased the chemosensor sensitivity to the TATAAA analyte by over 4 times compared to that of the nonimprinted polymer. The herein-devised detection platform enabled the generation of a library of hexamer probes for typing the majority of SNP probes as well as studying a molecular mechanism of the complex transcription machinery, physics of single polymer molecules, and stable genetic nanomaterials.

Inherently Chiral Spider-Like Oligothiophenes.

The racemate of an inherently chiral "spider-like" octathiophene monomer T83 , in which chirality is generated by torsion in its backbone, was synthesized. The racemate was resolved into configurationally stable antipodes by HPLC on a chiral stationary phase. Electrooxidation of the enantiomers resulted in materials displaying high enantiorecognition ability towards the antipodes of some chiral probes. Moreover, the T83 racemate demonstrated great aptitude to stimulate formation of 3D rigid architectures if used as a cross-linking monomer for molecular imprinting. This feature was exploited to devise a molecularly imprinted polymer-based chemosensor selective for a thymine-adenine oligonucleotide.

Molecularly imprinted polymers for separating and sensing of macromolecular compounds and microorganisms.

The present review article focuses on gathering, summarizing, and critically evaluating the results of the last decade on separating and sensing macromolecular compounds and microorganisms with the use of molecularly imprinted polymer (MIP) synthetic receptors. Macromolecules play an important role in biology and are termed that way to contrast them from micromolecules. The former are large and complex molecules with relatively high molecular weights. The article mainly considers chemical sensing of deoxyribonucleic acids (DNAs), proteins and protein fragments as well as sugars and oligosaccharides. Moreover, it briefly discusses fabrication of chemosensors for determination of bacteria and viruses that can ultimately be considered as extremely large macromolecules.

Electrochemically synthesized molecularly imprinted polymer of thiophene derivatives for flow-injection analysis determination of adenosine-5'-triphosphate (ATP).

Two selective chemosensors for adenosine-5'-triphosphate (ATP) determination featuring molecularly imprinted polymer (MIP) film recognition units were fabricated. For imprinting, three different thiophene derivatives were used as functional monomers. That is, the uracil substituent of bis(2,2'-bithienyl)methane 2 complementarily H-bond paired the adenine moiety of ATP, the boronic acid substituent of thiophene 3 covalently bound vicinal diols of the ribofuranose moiety, and amide substituents of bis(2,2'-bithienyl)methanes 4 bound to the pyrophosphate moieties. Different binding motifs adopted for the ATP recognition and the structure of the supramolecular pre-polymerization complex were optimized with the DFT computing at the B3LYP/3-21G((*)) level. MIP films were prepared by potentiodynamic electropolymerization of this complex with the imprinting factor of 9.47±0.2. An analytical signal was transduced with a 10-MHz resonator of EQCM and a Pt electrode for the piezoelectric microgravimetry (PM) and capacitive impedometry (CI) determination of ATP, respectively, under FIA conditions. Analytical properties of the MIP film were unraveled by spectroscopic ellipsometry, XPS, IRRAS, and DPV. The limit of detection was 0.1 and 0.2 µM for the PM and CI chemosensor, respectively, being an order of magnitude lower than the ATP concentration in biological systems. Moreover, cross-selectivity was demonstrated with the adenosine-5'-diphosphate (ADP) imprinting and ATP discrimination.

Electrochemically synthesized polymers in molecular imprinting for chemical sensing.

This critical review describes a class of polymers prepared by electrochemical polymerization that employs the concept of molecular imprinting for chemical sensing. The principal focus is on both conducting and nonconducting polymers prepared by electropolymerization of electroactive functional monomers, such as pristine and derivatized pyrrole, aminophenylboronic acid, thiophene, porphyrin, aniline, phenylenediamine, phenol, and thiophenol. A critical evaluation of the literature on electrosynthesized molecularly imprinted polymers (MIPs) applied as recognition elements of chemical sensors is presented. The aim of this review is to highlight recent achievements in analytical applications of these MIPs, including present strategies of determination of different analytes as well as identification and solutions for problems encountered.