Unveiling Secondary Alcohol Stereochemistry Through Experiments

Determining the absolute configuration of secondary alcohols is crucial to understanding the properties and functions of chiral molecules. While several methods are available for this process, the choice of method depends on the specific molecule and the resources available. One widely used technique is proton nuclear magnetic resonance (1H NMR) anisotropy, which is convenient as it does not require compound crystallization. Another method is thin-layer chromatography (TLC), which is cost-effective and sensitive. Other approaches include vibrational circular dichroism (VCD), Raman optical activity (ROA), and X-ray crystallography. The Mosher ester method, which involves the use of Mosher's acid, is also commonly employed.

Thin-layer chromatography

To begin, parallel reactions are set up for the secondary alcohols under study. These reactions utilise both enantiomers of a chiral kinetic resolution reagent, with one of the enantiomers being the fast-reacting reagent. The reactions are typically carried out at room temperature for about an hour, using small quantities of solvent, substrate, and catalyst. Deuterated chloroform is often used as the solvent to enable a comparative quantitative analysis of reaction conversion via 1H NMR spectroscopy.

After the reactions are complete, TLC is used to analyse the products. The TLC plate is spotted with an authentic sample of the compound, or standard, alongside the compound in question. The Rf value, which indicates the distance travelled on the TLC plate, can help identify the compound. If two substances have the same Rf value, they are likely the same compound, although this is not always the case.

Additionally, software such as ImageJ can be used to analyse the TLC plate and provide a quantitative analysis of reaction conversion. This technique is simple and effective, with a high success rate among students in determining the absolute configuration of unknown secondary alcohols.

Proton nuclear magnetic resonance (1H NMR) anisotropy method

Proton nuclear magnetic resonance (1H NMR) anisotropy is a widely used method for determining the absolute configuration of secondary alcohols. This technique is advantageous as it does not require the crystallization of compounds, and NMR machines are readily available. The 1H NMR anisotropy method is based on observing the anisotropy effect, which allows for the determination of the absolute configuration of the alcohol part.

One application of the 1H NMR anisotropy method is the Mosher method, also known as the advanced Mosher method, which involves the use of chiral auxiliaries such as Mosher's reagent (α-methoxy-α-(trifluoromethyl)phenylacetic acid or MTPA) and Trost's reagent (α-methoxyphenylacetic acid or MPA). The absolute configurations of these chiral auxiliaries are known, and the method focuses on rationalizing the preferred conformation of the esters formed with chiral secondary alcohols and MTPA or MPA. Additionally, the aromatic substituent (phenyl group) in this method generates a magnetic anisotropy effect due to the ring current induced under an external magnetic field.

Another technique that utilizes 1H NMR anisotropy is the fucofuranoside method, which employs 1H and 13C NMR spectra simultaneously. This method takes advantage of the strong anisotropy of pyridine, the solvent that solvates the polar substituent, instead of relying on the direct anisotropy of the substituent. The β-d- and β-l-fucofuranosyl groups, acting as polar and chiral substituents, provide an orientation effect (exo-anomeric effect) on the aglycon. This results in a more rigid conformation compared to other methods.

Furthermore, the Competing Enantioselective Conversion (CEC) method, in conjunction with thin-layer chromatography, can determine the absolute configuration of enantioenriched secondary alcohols. This approach requires micromole quantities of the secondary alcohol being tested and takes approximately 60 minutes. 1H NMR spectroscopy is then conducted on the samples to provide evidence of reaction conversion, supporting the qualitative nature of the CEC method.

In addition to these methods, it is worth noting that other techniques, such as vibrational circular dichroism (VCD) and Raman optical activity (ROA), can also be employed to determine absolute configuration. These methods involve comparing calculated and experimental spectra to assign the absolute configuration.

Vibrational circular dichroism (VCD)

VCD was first demonstrated in 1975, and the first VCD instrument was developed in 1973. However, it only started to gain popularity in the 21st century, and commercial instruments have been available since 1997. VCD is a widely used standard method for determining absolute stereochemistry and, to a lesser extent, biomolecule characterization and dynamic processes. It has been used extensively for both polypeptides and proteins in solution, with improved results over those obtained by visible/UV circular dichroism (CD) or optical rotatory dispersion (ORD).

In VCD, the output beam is often collimated or weakly focused to reduce artifacts. It then passes through a polarizer and a photo-elastic modulator (PEM), typically made of ZnSe, before being directed onto a sample. The beam is then focused onto a detector, usually with a ZnSe lens. The linear polarizer is positioned at a 45-degree angle with respect to the ZnSe crystal axis. The polarized radiation is doubly modulated, both by the PEM and the interferometer setup.

VCD is a valuable technique for determining the absolute configuration of organic molecules. It is a more general approach than other methods as it observes the vibration modes of atoms and is, therefore, applicable to most organic molecules. The absolute configuration is determined by comparing the experimental spectrum with a spectrum generated from quantum mechanical calculations. Modern DFT programs allow for reasonable predictions of VCD spectra, and the quality of these calculations can be assessed by comparing the calculated and measured IR spectra.

X-ray crystallography

The process of X-ray crystallography can be challenging, as it requires the growth of suitable crystals, which can sometimes take years. Additionally, heavy atoms, such as sulfur or chlorine, were traditionally needed to detect a strong enough anomalous signal. However, with recent advancements, such as the Hooft-Spek approach and improved detectors, it is now possible to determine absolute configurations with lighter atoms like oxygen.

In the context of secondary alcohols, X-ray crystallography can be employed after introducing a chiral auxiliary with a known absolute configuration. For example, (1S,2R,4R)-(−)- camphorsultam has been used as a chiral auxiliary for enantioresolving racemic carboxylic acids and determining their absolute configurations by X-ray crystallography. This methodology can be further extended by using chiral acids, such as camphorsultam dichlorophthalic acid (CSDP acid) and 2-methoxy-2-(1-naphthyl)propionic acid (MαNP acid), which are effective for enantioresolution of racemic alcohols and simultaneous determination of their absolute configurations.

The use of X-ray crystallography provides a direct experimental method for determining the absolute configuration of molecules, specifically the spatial arrangement denoted by R or S. While it may require significant resources and time, it offers a reliable approach for understanding the stereochemistry of compounds, which is crucial in fields such as drug discovery and development.

Competing Enantioselective Acylation (CEA)

The CEA method is innovative because it relies on differential reaction rates with enantioselective catalysts for optically enriched substrates. The rapid reaction kinetics are quantitatively estimated using LC/MS analysis as the characterization technique for the enantioselective transformations. This technique is used to compare the reaction rates of parallel reactions, acting as an indicator to prove enantioselective conversions.

The CEA reaction was first used to determine the absolute configuration at C-2 of a compound in a mixture. The acylation of the secondary alcohol at C-2 was performed using an enantiomeric pair of HBTM catalysts, allowing the reaction rates of parallel reactions to be compared using LC/MS analysis. This method was successful in determining the configuration, even though the compounds were obtained as a mixture.

The success of the CEA method demonstrates its practicality and simplicity, offering an efficient and time-effective approach for analyzing experimental results.

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