If you have a fairly basic commercial capillary electrophoresis (CE) system, with a fluorescence detector that can only detect a single wavelength at a time, how do you make it as sensitive as possible? Well, perhaps the only way is to optimize all the relevant separation parameters, such as buffer composition, pH level and applied voltage. And even after doing that, you’re still hampered by the fact your detector can only detect a single wavelength at a time, when the analytes in your sample will probably fluoresce over a range of different wavelengths.
Now, a team of Spanish chemists led by Ángel Ríos at the University of Castilla-La Mancha in Ciudad Real has come up with another way to enhance the sensitivity of basic CE systems, which also handily deals with the wavelength problem. And it’s as simple as adding a sprinkling of graphene quantum dots.
Graphene is a one-atom-thick sheet of carbon, famed for its immense strength and impressive conductivity. Exposing these sheets to strong acids causes them to break apart into small fragments, which sometimes then bind with each other. The end result is lots of small graphene particles, made up of up to 10 layers and around 10–30nm in size.
Termed graphene quantum dots, these particles possess various interesting electrical and optical properties, including the ability to fluoresce. Unlike many fluorescent compounds, however, they always fluoresce at the same main wavelength, irrespective of the wavelength used to excite them. This potentially makes them ideal for use with fluorescence detectors that can only detect a single wavelength at a time.
So, Ríos and his colleagues decided to see what happened when they mixed graphene quantum dots with several model analytes, comprising solutions of three different fluoroquinolone antibiotics: lomefloxacin, norfloxacin and ofloxacin. What they found was that not only did the graphene dots enhance the fluorescence of all three solutions, but the maximum emission for the three solutions was now at the same main wavelength, even though the antibiotics all fluoresce at slightly different wavelengths.
Further investigations confirmed that the negatively charged graphene quantum dots were binding with the three positively charged antibiotics, forming complexes that all fluoresced at the same main wavelength. Rather than the quantum dots staying independent and merely swamping the antibiotics’ fluorescence with their own.
Ofloxacin in milk
With this confirmation, Ríos and his colleagues then decided to see whether graphene quantum dots could help to detect ofloxacin, which is regularly used for treating livestock and is known to leave residues in tissues, in cow’s milk. For this, they used an inexpensive, commercially available CE system with a fluorescence detector that could only detect a single wavelength at a time.
After optimizing the separation parameters, they experimented with injecting the graphene quantum dots into the capillary both before and after the milk sample. This revealed that the quantum dots would only bind with the ofloxacin and enhance the fluorescence if they were added before the sample.
The scientists spiked the milk samples with ofloxacin at various concentrations, as the samples did not naturally contain any ofloxacin. They then found that enhancing the fluorescence with graphene quantum dots allowed them to detect the antibiotic at low nanogram per milliliter concentrations. This makes their method around 40 times more sensitive than a previous CE method that employed solid-phase extraction and around as sensitive as high-performance liquid chromatography, which requires much more expensive instrumentation. It is also faster than both these other methods, taking less than 15 minutes.
Ríos and his colleagues are now investigating what other types of CE analyses could benefit from a simple sprinkling of graphene quantum dots.