Ultrasensitive and label-free chirality detection of diabetes-related metabolic molecules

(Nanowerk Spotlight) Chirality is a critical concept in chemistry and life sciences, especially when applied to the molecular level. Many molecules such as amino acids, proteins, sugars, and DNA are chiral. Two mirror images of a chiral molecule are called enantiomers or optical isomers. Pairs of enantiomers are often designated as right-handed and left-handed.
"An abnormal concentration of chiral molecules has been observed in humans with increasing age and various chronic diseases, such as cancer, diabetes, kidney disease, and neurological disease, indicating the potential of applying chiral biomarkers as health indicators for diagnostic and prognostic applications," Yuebing Zheng, Associate Professor & William W. Hagerty Fellowship in Engineering at The University of Texas at Austin, tells Nanowerk. "In particular, elevated levels of many D-type metabolic molecules in urine have shown a strong correlation with diabetes mellitus."
The diabetes-induced change in chirality of urine metabolites has not been fully explored, hindering clinical development of the chirality-based disease diagnosis and monitoring. In particular, establishment of an accurate relationship between diabetes and chirality of metabolites in urine is crucial to improving the knowledge on the pathological roles of chiral disorder, which will facilitate new diagnostic device development.
So far, though, it has been very challenging to rapidly determine the chirality of urine metabolites with high accuracy.
A new method, developed by Zheng's group and reported in ACS Nano ("Label-Free Ultrasensitive Detection of Abnormal Chiral Metabolites in Diabetes"), can overcome the challenges in chiral sensing of metabolites by using microbubble-induced intense accumulation of biomolecules onto plasmonic chiral metamaterials with high sensitivity (100 pM) and low volume (10 µL).
"We have demonstrated label-free chiral detection of metabolic molecules at picomolar level through microbubble-induced rapid accumulation of biomolecules on plasmonic chiral sensors, which shows a 10-million times enhancement in sensitivity comparing to state-of-the-art plasmonic chiral sensors," says Yaoran Liu, a graduate student in Zheng's group and the paper's first author. "With its ultrahigh sensitivity, our technique uncovers the typically undetectable diabetes-induced abnormal dextrorotatory shift in total chirality of urine metabolites."
The team shows that monitoring such abnormal shifts of total chirality enables a diagnostic accuracy of 84%, a large improvement compared to the 72% for conventional glucose tests on clinical urine samples.
Schematic illustration of the collection and purification of urine samples, and the microbubble enabled accumulation of chiral metabolic molecules on chiral metasurface for enhanced chiral sensing and diabetic detection
a) Schematic illustration of the collection and purification of urine samples, and the microbubble enabled accumulation of chiral metabolic molecules on chiral metasurface for enhanced chiral sensing and diabetic detection; b) Normalized dissymmetry factors (ΔΔλ/λsum) measured for urine from normal and diabetic human; c) Receiver operating characteristic curves (ROC) of ΔΔλ/λsum and glucose concentration. (Image: Zheng research group, The University of Texas at Austin) (click on image to enlarge)
"So far, conventional chiroptical methods have suffered from large sample consumption and low molar sensitivity for metabolic molecules with ultra-small molecular mass and weak light-matter interactions, hindering their applications on detecting the trace chiral metabolites in urine," Liu points out.
Even though plasmon-enhanced chiral sensors can discriminate chiral molecules at picogram level with a wide range of molecular weights, the lowest detectable analyte concentration has been limited at ∼1 mM to ensure sufficient molecule-metamaterial interactions, hindering the chiral sensing of trace urine metabolomes in clinical applications.
In addition, plasmonic chiral sensing requires the analytes to be physically adsorbed on the plasmonic surfaces or residing near the superchiral fields with short (i.e. nanometer-scale) working distances.
The researchers achieved their ultrahigh sensitivity in chiral sensing of biomolecules by utilizing two enhancement mechanisms: the microbubble-induced accumulation of biomolecules onto the chiral plasmonic substrates; and the subsequent plasmon-enhanced chiral sensing.
The plasmonic substrate used is based on previous work by Zheng's group on plasmonic moiré chiral metamaterials (read our previous Nanowerk Spotlight on this work: "A new type of ultra-thin plasmonic chiral metamaterial"). In the present work, they apply this metamaterial – which consist of two layers of gold nanohole arrays stacked into moiré patterns – to generate both the optothermal microbubbles and the superchiral fields.
Liu explains the working principle of this technique: "The irradiation of a focused laser onto the metamaterial induces plasmon-enhanced optical heating at the laser focus point, vaporizing the solution above the substrate and generating a microbubble. The microbubble-induced Marangoni convection can effectively drag biomolecules in the solution toward the laser spot."
"The increased concentration of molecules near the substrate – in an area measuring approx. 5 µm2 – and the strong downward forces at the stagnation area near the microbubble-substrate interfaces then effectively print the molecules onto the plasmonic substrate with high binding affinity, enabling effective molecule accumulation for enhanced sensitivity" he adds.
The team is currently working on enhancing the specificity via improved filtering or integration of microfluidic-based separation techniques. They are also applying this technique for prognosis of chronic kidney disease.
"The accumulation-assisted plasmonic chiral sensing shows great potential in development of point-of-care devices for first-line noninvasive screening and prognosis of early-stage pre-diabetes or diabetes and its complications," Zheng concludes. "As chiral molecules have been found to be altered in several chronic disease such as atherosclerosis, neurodegenerative diseases and cancers as well, we envision a routine chiral detection that is sensitive and non-invasive could also be used as new tools for early chronic disease detection."
The main issue in the team's current technology is the separation of the molecules of interests from the complex biological solution. However, they are confident that combining the current chiral detection technology with multiplexing chiral separation module will potentially allow the specific detection of trace level chiral biomarker in complex human biofluidic for disease detection application.
By Michael is author of three books by the Royal Society of Chemistry:
Nano-Society: Pushing the Boundaries of Technology,
Nanotechnology: The Future is Tiny, and
Nanoengineering: The Skills and Tools Making Technology Invisible
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