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Separating Left and Right: A New Way to Differentiate Chiral Compounds

Figure Created in Biorender

Writer: Matthew Liu ‘26

Editor: Yilin Xie ‘26

Whenever you have a headache and reach for that bottle of ibuprofen, chances are that you are taking one out of two possible versions of that compound. The benefits of ibuprofen come exclusively from the left-handed version while the other does not contribute at all [1]. Ibuprofen is far from the only pharmaceutical where the handedness matters for maximizing efficacy while minimizing side effects: over 50% of drugs currently produced are chiral compounds [2, 3]. Imagine your left and right hands: they are mirror images of each other, but you cannot impose one onto the other. This is what chirality refers to: a molecule that can occur in two asymmetric forms that are non-superimposable mirror images of each other, and the two versions are referred to as enantiomers [4]. As a result, reliable methods to isolate pure chiral compounds are important for drug development. This year, researchers at Tsinghua University developed a novel method to distinguish different enantiomers of compounds, using mass spectrometry [5].

The differences between enantiomers may not seem like much at first, but they are pretty important for biological processes. Enantiomers usually have identical physical properties except when they interact with other chiral compounds, in which there can be different outcomes of reactivity. To denote the handedness of an enantiomer, chemists usually designate a unique symbol for each of the two configurations. In nature, chiral amino acids are found as “left-handed” enantiomers while carbohydrates exist mainly as “right-handed” enantiomers [6]. Since our bodies are composed of amino acids and carbohydrates, they naturally prefer a certain chirality; in the context of pharmaceuticals, the wrong enantiomer can have no or even deadly effects in some instances. For example, R-albuterol helps with difficulty breathing by relaxing the lungs while S-albuterol constricts airways and induces many inflammatory and detrimental effects on the lungs; R-thalidomide is an effective sedative whereas S-thalidomide causes more than 10,000 newborn birth defects [6,7]. These potential risks make it essential that potential clinical candidates have been correctly identified and are free of impurities and variants. However, there are some drugs that cannot be commercially available because separating the chiral forms has been difficult or production costs are too high [3]. 

Currently, the most commonly used methods for chiral separation of pharmaceuticals are high-performance liquid chromatography, organic solvent chromatography, and supercritical fluid chromatography [8-11]. While these chromatographic methods are effective at controlling enantiomeric purity, they may be very time-consuming, and the reaction conditions for separation must be tailored to the unique properties of each compound, which may be unknown [3]. In the age of computational methods, virtual screening and molecular modeling have been proposed as alternatives due to their rapid identification of potential candidates, but reliable calculations rely on an accurate depiction of the stereochemistry of each enantiomer since they have unique binding modes, which requires having a pure sample of that specific chiral compound [4]. Enter mass spectrometry (MS), which has a short analysis time, a low limit of compound detection, and a great ability to detect selective compounds from a complex mixture without interference. The major obstacle is that MS intrinsically is chiral-blind because two enantiomers typically have the same mass and spectra, so a chiral selector would need to be injected into the solution [12]. The only methods available have been using a chiral gas to induce collisions and dissociations, which takes advantage of the fact that dissociation rates are different for enantiomers. Otherwise, MS has been under-exploited yet has more potential, which is what motivated the researchers at Tsinghua University to develop their new technique that does not rely on a chiral gas reference [6]. 

The researchers were able to differentiate enantiomers by placing the compounds in an electromagnetic field that was created using two alternating current sources that allowed for separate excitation. Because of the asymmetrical nature of the enantiomers, there is a slight polarity difference between them, and the electromagnetic field induces rotational trajectories in each enantiomer that are opposite of each other. These different rotations change the amount of drag force each enantiomer experiences when they collide with background gas molecules, creating a significant separation in the macro motions. The differences in collision properties, though small, are significant enough to allow super high-resolution separation and analysis of the ejected ions. The method utilized an ion trap MS (IM-MS), which is also used for structural analysis, so not only can IM-MS separate the enantiomers, but it can also identify their structural properties. The researchers showed the applicability of this technique to various chiral macromolecules and enantiomers with multiple stereocenters, including amino acids, carbohydrates, metabolite compounds, and even current small-molecule drugs. The run time often lasted less than a minute, proving it a fast and facile means for identifying and separating the chirality of candidates for drug discovery and from synthesis. Moreover, the technique allowed the determination of enantiomeric ratios in the mixture, bringing further applications as enantiomeric ratios can be an indication of the cause of diseases [6].

Overall, using directional rotations of ions in MS has shown promising potential to be a quicker, more efficient way to purify stereoselective compounds. The fact that the researchers did not need to use any chiral references and simply implemented an ion trap is interesting in that a chiral-blind method became chiral-sensitive using electromagnetic properties. Moreover, it highlights the readily available use of small instruments and simple procedures that we may be under-utilizing: with a few tweaks, we could make them viable for many other purposes. While they did not test diastereomers or determine absolute chiral configurations solely based on this method, the new MS system could be scaled up and become a new conventional procedure to mass-produce one enantiomer of therapeutic compounds. As a result, we could potentially see a new era of innovation and advances that produce new cures and reshape our health and lifestyles.


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