Optical sensing of chiral alcohols including cryptochiral alcohols displaying -a-, b-,y-,and o-stereocenters or chirality by isotopic substitution

Abstract

The present disclosure is directed to an analytical method for determining the concentration of a chiral alcohol in a sample and one or both of (i) the absolute configuration of the chiral alcohol in the sample, and (ii) the enantiomeric and / or the diastereomeric composition of the chiral alcohol in the sample. The present disclosure is also directed to an aryl or heteroaryl sulfonamide probe having the structure according to formula (I): Formula (I), where Ar, R1, R2, and n are as described herein, as well as to a kit for carrying out the analytical method described herein.

Description

– 1 – OPTICAL SENSING OF CHIRAL ALCOHOLS INCLUDING CRYPTOCHIRAL ALCOHOLS DISPLAYING α-, β-, γ- AND δ-STEREOCENTERS OR CHIRALITY BY VIRTUE OF ISOTOPIC SUBSTITUTION

[0001] This application claims the priority benefit of U.S. Provisional Application Serial No. 5 63 / 660,163, filed June 14, 2024, which is hereby incorporated by reference in its entirety.

[0002] This invention was made with government support under CHE-2246747 awarded by the National Science Foundation. The government has certain rights in the invention. FIELD 10

[0003] The present disclosure relates to an analytical method for the determination of the concentration and one or both of the absolute configuration and the enantiomeric and / or the diastereomeric composition of an analyte in a sample, based on optical sensing. BACKGROUND

[0004] The Mitsunobu reaction is among the most frequently used transformations in 15 asymmetric synthesis (Swamy et al., Chem. Rev.109:2551-2651 (2009); Fletcher, Org. Chem. Front.2:739-752 (2015); Munawar et al., Molecules 27:6953 (2022)). It achieves high-yielding stereospecific conversion of a chiral alcohol to a new functional group with a variety of pronucleophiles in the presence of a phosphine and an azodicarboxylate. These readily available reagents work in tandem to generate a free nucleophile and a reactive alkoxyphosphonium salt, 20 thus setting the stage for efficient one-pot functional group modification under mild conditions. The outstanding operational simplicity, click chemistry-like features and reliable inversion of the absolute configuration at the chiral carbon center have been instrumental in literally countless synthetic applications, in particular in total natural product synthesis. Since its introduction more than half a century ago (Mitsunobu et al., Bull. Chem. Soc. Jpn.40:935-939 (1967); Mitsunobu 25 and Yamada, Bull. Chem. Soc. Jpn.40:2380-2382 (1967)), the steadily increasing significance and popularity of the Mitsunobu reaction have inspired several mechanistic investigations (Ahn et al., J. Org. Chem.67:1751-1753 (2002); Schenk et al., J. Am. Chem. Soc.127:12566-12576 (2005)) as well as innovative modifications and developments aimed at improving the synthetic value and scope (But and Toy, J. Am. Chem. Soc.128:9636–9637 (2006); Hirose et al., Angew. 30 Chem. Int. Ed.52:4613-4617 (2013); Buonomo and Aldrich, Angew. Chem. Int. Ed.54:13041- 13044 (2015); Guo et al., Angew. Chem. Int. Ed.63:e202402878 (2024)). By contrast, other potential uses of this powerful reaction have been largely neglected to date. 310804994v3– 2 –

[0005] Optical analysis of the amount and enantiomeric composition of chiral compounds performed directly in mixtures without cumbersome chromatographic separation or other tedious work-up has received increasing attention in recent years (Pu, Chem. Rev.104:1687-1716 (2004); Leung et al., Chem. Soc. Rev.41:448-479 (2012);Wolf and Bentley, Chem. Soc. Rev. 5 42:5408-5424 (2013), Herrera et al., J. Am. Chem. Soc.140:10385-10401 (2018); Quan et al., Angew. Chem. Int. Ed.61:e202201258 (2022); Formen et al., Angew. Chem. Int. Ed. 63:e202400767 (2024), Herrera et al., J. Am. Chem. Soc.141:11151-11160 (2019); De los Santos et al., Angew. Chem. Int. Ed.59:2440-2448 (2020); Sasaki et al., Chem. Sci.11:3790- 3796 (2020); Wang et al., J. Am. Chem. Soc.139:8436-8439 (2017); Hassan and Wolf, Nat. 10 Comm.12:6451 (2021)). This field has important applications across the chemical and health sciences, for example chromatography-free asymmetric reaction screening with crude mixtures or biomarker analysis, and bears huge promise to streamline numerous chiral compound development projects (Hassan et al., Synthesis 54:2527-2538 (2022)). The state of the art of chirality sensing, however, is mostly confined to opportune compound classes like amines, 15 amino alcohols, amino acids and α-hydroxy acids while alcohols have remained difficult (Dragna et al., J. Am. Chem. Soc.134:4398-4407 (2012); Wezenberg et al., Angew. Chem., Int. Ed. 50:713-716 (2011); Kim et al., Angew. Chem. Int. Ed.47:8657-8660 (2008); Biedermann and Nau, Angew. Chem. Int. Ed.53:5694-5699 (2014); Wang et al., Angew. Chem. Int. Ed. 61:e202211853 (2022); Wang et al., Angew. Chem. Int. Ed.59:23817–23824 (2020); Pilicer et 20 al., J. Am. Chem. Soc.139:1758-1761 (2017); Joyce et al., Chem. Sci.5:2855-2861 (2014); De los Santos et al., Angew. Chem. Int. Ed.58:1198-1202 (2019); Badetti et al., Chem. Eur. J. 22:6515-6518 (2016); Zardi et al., J. Am. Chem. Soc.139:15616-15619 (2017); Zhu et al., Angew. Chem. Int. Ed.59:10868–10872 (2020); Shirbhate et al., J. Am. Chem. Soc.142:4975- 4979 (2020); Formen and Wolf, Angew. Chem. Int. Ed.60:27031-27038 (2021); Li, et al., Chem. 25 Sci.12:2504-2508 (2021); Howard et al., J. Am. Chem. Soc.144:17269–17276 (2022)).

[0006] In most cases, chiral compounds exhibit weak, if any, blue-shifted circular dichroism (CD), UV or fluorescence signals that are insufficient for quantitative sensing purposes. This shortcoming can be addressed by introducing chromophoric probes through covalent bond formation or other means. The inherently weak nucleophilicity of alcohols, however, continues 30 to hamper progress in this field and a practical method that is robust and broadly applicable but avoids harsh conditions, the use of unstable reagents, equilibria prone to side reactions and complicated sample processing is still elusive despite remarkable improvements in recent years (You et al., Nat. Chem.3:943–948 (2011); You et al., J. Am. Chem. Soc.134:7117–7125 (2012); Ni et al., Angew. Chem. Int. Ed.57:1300–1305 (2018); Thanzeel et al., Angew. Chem. Int. Ed. 310804994v3– 3 – 59:21382–21386 (2020); Nelson et al., Chem. Sci.12:8784-8790 (2021); De los Santos et al., Chem. Eur. J.28:e202202028 (2022)).

[0007] The present application is directed to overcoming these and other deficiencies in the art. 5 SUMMARY

[0008] A first aspect of the present application relates to an analytical method that includes the steps of: providing a sample potentially containing a chiral alcohol that can exist in stereoisomeric forms; providing an aryl or heteroaryl sulfonamide probe; contacting the sample with the probe, wherein said contacting is carried out under conditions to permit reaction of the 10 probe with the chiral alcohol, if present in the sample, to form a probe-labeled analyte; and detecting the probe-labeled analyte in the sample using one or more chiroptical assay formats, and determining the concentration of the chiral alcohol in the sample and one or both of (i) the absolute configuration of the chiral alcohol in the sample, and (ii) the enantiomeric and / or the diastereomeric composition of the chiral alcohol in the sample. 15

[0009] A second aspect of the present application relates to an aryl or heteroaryl sulfonamide probe having the structure according to formula (I):wherein Ar is a substituted or unsubstituted aromatic or heteroaromatic group, R1is H, a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl, 20 R2is a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl, and n is 0, 1, 2, 3, 4, or 5.

[0010] In certain embodiments, the aryl or heteroaryl sulfonamide probe having the structure according to formula (I) is used in carrying out the method according to the first aspect of the present application. 25

[0011] A third aspect of the present application relates to a kit for carrying out the analytical method described herein. The kit may include a solution comprising an aryl or heteroaryl sulfonamide probe according to the second aspect of the present application and, optionally, one or more of (i) sample tubes suitable for use with a spectrophotometer; (ii) an optically pure reference sample of a chiral alcohol; (iii) directions for using a spectrophotometer for carrying 30 out circular dichroism (CD), vibrational CD (VCD), electronic CD, optical rotatory dispersion (ORD), or polarimetry analyses to measure the concentration of an analyte in a sample and one or both of the absolute configuration of the analyte in the sample, and the enantiomeric and / or 310804994v3– 4 – the diastereomeric composition of the analyte in the sample; and (iv) a recordable medium comprising a template for analyzing data obtained from the spectrophotometer and determining the concentration of an analyte in a sample and one or both of the absolute configuration of the analyte in the sample, and the enantiomeric and / or the diastereomeric composition of the analyte 5 in the sample.

[0012] As demonstrated in the accompanying Examples, some of the unique features of the Mitsunobu reaction can overcome issues associated with chiroptical sensing of chiral alcohols. To overcome the issues preventing the use of chromophobic probes covalently attached to alcohols, the Mitsunobu reaction has been repurposed and applied in a conceptually new sensing 10 approach that, for the first time, neither incorporates the alcohol into an acetal, carbamate or another functionality nor requires formation of a metal complex or supramolecular assembly, but instead installs irreversibly and enantiospecifically an optimized sulfonamide chromophore directly at the chiral carbon center (FIG.1). This new sensing strategy has several advantages. The ultimate contiguity of stereocenter and chromophore is conducive to strong CD inductions at 15 long wavelengths, which is a prerequisite for precise quantitative analysis and reduces the possibility of molecular interferences one might expect, for example, in multi-compound mixtures. Moreover, it enables sensing of very challenging chiral targets. Unmatched sensing scope and utility are demonstrated with alcohols having α-, β-, γ- or even δ-stereocenters and with an alcohol chiral by virtue of isotopic substitution. Notably, all these tasks become possible 20 with a single probe and a unique CD / UV relay assay that exploits the Mitsunobu triphenylphosphine reagent in two ways: it mediates the chromophore placement directly at the chiral center to induce (i) a remarkably strong, red-shifted CD signal that increases linearly with the sample er; and (ii) it also participates in a redox reaction with a quinone molecule to generate a characteristic UV response that is independent of the enantiopurity of the alcohol and thus25 allows determination of the total analyte concentration. This reaction sequence thus enables in- tandem determination of the enantiomeric ratio (er) and concentration of alcohol samples by using readily available chemicals that produce strong chiroptical signals at long wavelengths. The robustness of the reaction-based chiroptical assay is validated by accurate asymmetric reaction analysis with crude product mixtures. This eliminates the common need for 30 chromatographic work-up and outperforms traditional workflows with superior speed while reducing workload and chemical waste production. This is a highly sought-after goal in the pharmaceutical industry. 310804994v3– 5 – BRIEF DESCRIPTION OF THE DRAWINGS

[0013] FIG.1 is a schematic illustration showing chiroptical sensing challenges, advances, and applications.

[0014] FIGS.2A-2G illustrate assay components, chiral alcohol sensing concept, and 5 examples of CD and UV inductions. FIG.2A shows structures of the probes and initially tested alcohols (only one enantiomer is shown for simplicity). FIG.2B shows details of the CD / UV relay assay. FIG.2C shows representative CD spectra for S / R-11 and S / R-17 obtained at 1.7 mM in THF. FIG 2D shows linear CD amplitude increase as a function of sample %ee using probe 3 and alcohol 16. FIG.2E shows solvent screening of the CD sensing using alcohol 18 as test 10 analyte for probes 1, 3, 4, and 6. All spectra were collected at 1.7 mM. FIG.2F is a heat map showing the probe performances in six different solvents. The increase in the CD amplitude is visualized using lighter color (yellow or pale green in color version) to reflect low signal induction to darker color (dark green in color version) to reflect high signal induction. FIG.2G shows the UV change obtained by consumption of surplus PPh3with quinone 27 is directly 15 correlated to the initial alcohol concentration.

[0015] FIGS.3A-3B illustrate remote chirality sensing of chiral alcohols 30-35 with probe 3. FIG.3A shows the structures of the chiral alcohols. FIG.3B shows the CD spectra obtained at 1.7 mM using THF as the diluting solvent.

[0016] FIG.4 illustrates the CD sensing of the enantiomers of 36 and the structures of 38 and 20 39. The CD spectra were obtained at 1.7 mM using THF as the diluting solvent.

[0017] FIGS.5A-5B illustrate the asymmetric reaction analysis using traditional methods and chiroptical sensing. In FIG.5A, autocatalytic asymmetric ketone reduction of chiral ketone 40 is shown, yielding alcohol 11. In FIG.5B, a comparison of traditional and chiroptical sensing approaches is illustrated. AC=Absolute configuration. Conv.=Conversion. 25

[0018] FIG.6 illustrates the reaction scheme for the chiroptical sensing of (S)-1-methoxy-2- propanol with sulfonamide probes 1-10.

[0019] FIG.7 illustrates the results of chiroptical sensing of (S)-1-methoxy-2-propanol with sulfonamide probes 1-10. CD measurements were taken at 2.0 mM in CH2Cl2.

[0020] FIG.8 illustrates NMR analysis of the reaction between 3 and (S)-1-phenylethanol. 30

[0021] FIG.9 illustrates NMR analysis of the reaction between 1 and (S)-1-phenylethanol.

[0022] FIG.10 illustrates CD spectra of the sensing reaction of (S)-1-methoxy-2-propanol with probe 3 in CH3CN: CH2Cl2(3:1, v / v) followed by dilution in respective solvent. CD measurements were taken at 1.70 mM. 310804994v3– 6 –

[0023] FIG.11 illustrates CD spectra of the sensing reaction of (S)-1-methoxy-2-propanol with probe 1 in CH3CN: CH2Cl2(3:1, v / v) followed by dilution in respective solvent. CD measurements were taken at 1.70 mM.

[0024] FIG.12 illustrates CD spectra of the sensing reaction of (S)-1-methoxy-2-propanol 5 with probe 4 in CH3CN: CH2Cl2(3:1, v / v) followed by dilution in respective solvent. CD measurements were taken at 1.70 mM.

[0025] FIG.13 illustrates CD spectra of the sensing reaction of (S)-1-methoxy-2-propanol with probe 6 in CH3CN: CH2Cl2(3:1, v / v) followed by dilution in respective solvent. CD measurements were taken at 1.70 mM. 10

[0026] FIG.14 illustrates stereoinversion of alcohol 11 to the corresponding sulfonamide 43.

[0027] FIG.15 illustrates the crystal structure of (S)-N-(1-phenylethyl)-N- (phenylsulfonyl)benzenesulfonamide.

[0028] FIG.16 illustrates the reaction of 27 with PPh3. 15

[0029] FIG.17 illustrates the change in the UV absorbance produced by the UV relay assay as a function of the initial 2-butanol concentration.

[0030] FIG.18 illustrates MS spectrum of the reaction products obtained from 27 and PPh3(top spectrum). Zoomed in view of the signal assigned to 28 (middle spectrum). Zoomed in view of the signal assigned to 29 with the characteristic M+2 chlorine isotope pattern (bottom 20 spectrum).

[0031] FIG.19 illustrates the reaction of probe 3 with alcohols 11-22 and 30-36.

[0032] FIG.20 illustrates CD spectra obtained by applying probe 3 to (R)-11 (blue in color version) and (S)-11 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0033] FIG.21 illustrates CD spectra obtained by applying probe 3 to (R)-12 (blue in color 25 version) and (S)-12 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0034] FIG.22 illustrates CD spectra obtained by applying probe 3 to (R)-13 (blue in color version) and (S)-13 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0035] FIG.23 illustrates CD spectra obtained by applying probe 3 to (R)-14 (blue in color version) and (S)-14 (red in color version). CD measurements were taken at 1.2 mM in THF. 30

[0036] FIG.24 illustrates CD spectra obtained by applying probe 3 to (R)-15 (blue in color version) and (S)-15 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0037] FIG.25 illustrates CD spectra obtained by applying probe 3 to (R)-30 (blue in color version) and (S)-30 (red in color version). CD measurements were taken at 1.2 mM in THF. 310804994v3– 7 –

[0038] FIG.26 illustrates CD spectra obtained by applying probe 3 to (R)-31 (blue in color version) and (S)-31 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0039] FIG.27 illustrates CD spectra obtained by applying probe 3 to (R)-32 (blue in color version) and (S)-32 (red in color version). CD measurements were taken at 1.7 mM in THF. 5

[0040] FIG.28 illustrates CD spectra obtained by applying probe 3 to (R)-16 (blue in color version) and (S)-16 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0041] FIG.29 illustrates CD spectra obtained by applying probe 3 to (R)-17 (blue in color version) and (S)-17 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0042] FIG.30 illustrates CD spectra obtained by applying probe 3 to (R)-18 (blue in color 10 version) and (S)-18 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0043] FIG.31 illustrates CD spectra obtained by applying probe 3 to (R)-19 (blue in color version) and (S)-19 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0044] FIG.32 illustrates CD spectra obtained by applying probe 3 to (R,R)-20 (blue in color version) and (S,S)-20 (red in color version). CD measurements were taken at 1.2 mM in THF. 15

[0045] FIG.33 illustrates CD spectra obtained by applying probe 3 to (R)-21 (blue in color version) and (S)-21 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0046] FIG.34 illustrates CD spectra obtained by applying probe 3 to (R)-22 (blue in color version) and (S)-22 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0047] FIG.35 illustrates a CD spectrum obtained by applying probe 3 to (S)-33 (red in color 20 version). CD measurements were taken at 1.7 mM in THF.

[0048] FIG.36 illustrates a CD spectrum obtained by applying probe 3 to (S)-34 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0049] FIG.37 illustrates a CD spectrum obtained by applying probe 3 to (S)-35 (red in color version). CD measurements were taken at 1.7 mM in THF. 25

[0050] FIG.38 illustrates CD spectra obtained by applying probe 3 to (R)-36 (blue in color version) and (S)-36 (red in color version). CD measurements were taken at 1.7 mM in THF.

[0051] FIG.39 illustrates a CD spectrum obtained by applying probe 3 to cholesterol, 38. CD measurements were taken at 1.2 mM in THF.

[0052] FIG.40 illustrates a CD spectrum obtained by applying probe 3 to podophyllotoxin, 30 39. CD measurements were taken at 1.2 mM in THF.

[0053] FIG.41 illustrates the change in the UV absorbance produced by the UV relay assay as a function of the initial 2-butanol concentration. 310804994v3– 8 –

[0054] FIG.42 illustrates colorimetric change observed by the relay assay (remaining [PPh3] + quinone indicator). Concentration of remaining PPh3from left to right: 40.0 mM, 30.0 mM, 20.0 mM, 10.0 mM, 0.0 mM.

[0055] FIG.43 illustrates UV absorbance at 340 nm vs concentration of (S)-butanol. 5

[0056] FIG.44 illustrates chiroptical response of probe 3 to scalemic samples of 2-butanol.

[0057] FIG.45 illustrates CD amplitude at 325 nm vs sample ee.

[0058] FIG.46 illustrates autocatalytic reduction of acetophenone, and subsequent analysis via CD & UV spectroscopy and chiral GC-MS.

[0059] FIG.47 illustrates change in the UV absorbance produced by the UV relay assay as a 10 function of the initial phenylethan-1-ol concentration.

[0060] FIG.48 illustrates UV absorbance at 340 nm vs concentration of phenylethan-1-ol.

[0061] FIG.49 illustrates chiroptical response of probe 3 to scalemic samples of phenylethan-1-ol.

[0062] FIG.50 illustrates CD amplitude at 325 nm vs sample ee. 15

[0063] FIG.51 illustrates enantioseparation of racemic phenylethan-1-ol. Column: 2,6- dimethyl-3-pentyl-β-cyclodextrin (50% in polysiloxane) column, 100 °C, pressure: 12.9 psi Helium.

[0064] FIG.52 illustrates GC-MS of enantioenriched (S)-phenylethan-1-ol (Entry 1). Column: 2,6-dimethyl-3-pentyl-β-cyclodextrin (50% in polysiloxane) column, 100 °C, pressure: 20 12.9 psi Helium.

[0065] FIG.53 illustrates GC-MS of enantioenriched (S)-phenylethan-1-ol (Entry 2). Column: 2,6-dimethyl-3-pentyl-β-cyclodextrin (50% in polysiloxane) column, 100 °C, pressure: 12.9 psi Helium.

[0066] FIG.54 illustrates GC-MS of racemic phenylethan-1-ol (Entry 3). Column: 2,6- 25 dimethyl-3-pentyl-β-cyclodextrin (50% in polysiloxane) column, 100 °C, pressure: 12.9 psi Helium.

[0067] FIG.55 illustrates GC-MS of enantioenriched (S)-4-bromophenylmethan-d-ol. Column: 2,6-dimethyl-3-pentyl-β-cyclodextrin (50% in polysiloxane) column, 130 °C, pressure: 12.9 psi Helium. 30

[0068] FIG.56 illustrates GC-MS of enantioenriched (R)-4-bromophenylmethan-d-ol.

[0069] FIG.57 illustrates Mass spectrum of enantioenriched (S)-4-bromophenylmethan-d-ol. 310804994v3– 9 – DETAILED DESCRIPTION

[0070] One aspect of the present application relates to an analytical method for measuring the concentration of a chiral alcohol in a sample and one or both of the absolute configuration of the chiral alcohol in the sample, and the enantiomeric and / or the diastereomeric composition of 5 the chiral alcohol in the sample.

[0071] According to one embodiment, the analytical method includes the steps of: (1) providing a sample potentially containing a chiral alcohol that can exist in stereoisomeric forms; (2) providing an aryl or heteroaryl sulfonamide probe, preferably an achiral sulfonamide 10 according to formula (I) as disclosed herein; (3) contacting the sample with the probe, wherein said contacting is carried out under conditions to permit reaction of the probe with the chiral alcohol, if present in the sample, to form a probe-labeled analyte; and (4) detecting the probe-labeled analyte in the sample using one or more chiroptical assay 15 formats, and determining the concentration of the chiral alcohol in the sample and one or both of (i) the absolute configuration of the chiral alcohol in the sample, and (ii) the enantiomeric and / or the diastereomeric composition of the chiral alcohol in the sample.

[0072] A further aspect of the present application relates to the use of an aryl or heteroaryl 20 sulfonamide probe as defined below in an analytical method for measuring the concentration of an analyte in a sample and one or both of the absolute configuration of the analyte in the sample, and the enantiomeric and / or the diastereomeric composition of the analyte in the sample.

[0073] The probes of the present application comprise achiral, substituted or unsubstituted aryl or heteroaryl sulfonamides. Suitable probes can be achiral sulfonamide compounds having 25 the structure according to formula (I):(I), wherein: Ar is a substituted or unsubstituted aromatic or heteroaromatic group, R1is H, a substituted or unsubstituted aryl, or a substituted or unsubstituted 30 heteroaryl, R2is a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl, and 310804994v3– 10 – n is 0, 1, 2, 3, 4, or 5.

[0074] In some embodiments, the substituted or unsubstituted aryl at Ar, R1, and R2can be independently selected from the group consisting of phenyl, naphthyl, azulenyl, phenanthrenyl, anthracenyl, fluorenyl, pyrenyl, triphenylenyl, chrysenyl, naphthacenyl, biphenyl, triphenyl, and 5 tetraphenyl. In other embodiments, the substituted or unsubstituted heteroaryl at Ar, R1, and R2can be independently selected from the group consisting of pyrrolyl, pyrazolyl, imidazolyl, triazolyl, furyl, thiophenyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, triazinyl, thienopyrrolyl, furopyrrolyl, indolyl, azaindolyl, isoindolyl, indolinyl, indolizinyl, indazolyl, benzimidazolyl, imidazopyridinyl, 10 benzotriazolyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, pyrazolopyridinyl, triazolopyridinyl, thienopyridinyl, benzothiadiazolyl, benzofuyl, benzothiophenyl, quinolinyl, isoquinolinyl, tetrahydroquinolyl, tetrahydroisoquinolyl, cinnolinyl, quinazolinyl, quinolizilinyl, phthalazinyl, benzotriazinyl, chromenyl, naphthyridinyl, acrydinyl, phenanzinyl, phenothiazinyl, phenoxazinyl, pteridinyl, and purinyl. 15

[0075] According to one embodiment, Ar of formula (I) can bewherein: the squiggly line represents the point of attachment to the −S(O2)− group in formula I; each X is independently C or N, except that no more than three ring nitrogens are 20 present in the substituted or unsubstituted aromatic or heteroaromatic ring, and R3, R4, R5, R6, and R7are independently selected from the group consisting of a lone pair (when X is N), –H, –CN, –NO2, halogen, –C1-C6alkyl, –C1-C6alkoxy, –N–(alkyl)2, –C1-C6alkenyl, –C1-C6alkynyl, – C1-C6perfluoroalkyl, –aryl, –perfluoroaryl, –aryloxy, –N–(aryl)2, –heteroaryl, –O–heteroaryl, 25 –N–(heteroaryl)2, –cycloalkyl, –O–cycloalkyl, –N–(cycloalkyl)2, –heterocycloalkyl, –O–heterocycloalkyl, –N–(heterocycloalkyl)2, –C(O)Ra, –SO2Ra, and –OC(O)Ra; each Rais independently selected from the group consisting of –alkyl, –O–alkyl, –N–(alkyl)2, –alkenyl, –alkynyl, –aryl, –O–aryl, –N–(aryl)2, –heteroaryl, 310804994v3– 11 – –O–heteroaryl, –N–(heteroaryl)2, –cycloalkyl, –O–cycloalkyl, –N–(cycloalkyl)2, –heterocycloalkyl, –O–heterocycloalkyl, and –N–(heterocycloalkyl)2; and wherein, optionally, R3and R4, R4and R5, R5and R6, and / or R6and R7are alternatively taken together with the carbon or nitrogen atoms to which they are 5 attached to form a fused monocyclic or bicyclic ring system selected from the group consisting of cycloalkyl, heterocycloalkyl, aryl, and heteroaryl, wherein the fused ring system is optionally substituted with one or more groups selected from –alkyl, –O–alkyl, –N–(alkyl)2, –alkenyl, –alkynyl, –O-aryl, –O-heteroaryl, –N-(aryl)2, –N-(heteroaryl)2, –aryl, –C(O)Rc, –CO2Rb, –O-C(O)Rb, –NHC(O)Rb, –NRcC(O)Rb, 10 –NO2, –CN, –halogen, and –SO2Rb, wherein each Rbis independently aryl, heteroaryl, alkyl, or CH2Ar.

[0076] In certain embodiments, the probe Ar of formula I has one or two of R1, R2, R3, and R4that is –NO2or –OMe.

[0077] Exemplary probes are identified in the accompanying Examples and include, without 15 limitation:

[0078] As used herein, the term “alkyl” refers to a straight or branched, saturated aliphatic radical containing one to about twenty (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 20 18, 19, 20, 1–2, 1–3, 1–4, 1–5, 1–6, 1–7, 1–8, 1–9, 1–10, 1–11, 1–12, 1–13, 1–14, 1–15, 1–16, 1–17, 1–18, 1–19, 1–20, 2–3, 2–4, 2–5, 2–6, 2–7, 2–8, 2–9, 2–10, 2–11, 2–12, 2–13, 2–14, 2–15, 2–16, 2–17, 2–18, 2–19, 2–20, 3–4, 3–5, 3–6, 3–7, 3–8, 3–9, 3–10, 3–11, 3–12, 3–13, 3–14, 3– 15, 3–16, 3–17, 3–18, 3–19, 3–20, 4–5, 4–6, 4–7, 4–8, 4–9, 4–10, 4–11, 4–12, 4–13, 4–14, 4–15, 4–16, 4–17, 4–18, 4–19, 4–20, 5–6, 5–7, 5–8, 5–9, 5–10, 5–11, 5–12, 5–13, 5–14, 5–15, 5–16, 25 5–17, 5–18, 5–19, 5–20, 6–7, 6–8, 6–9, 6–10, 6–11, 6–12, 6–13, 6–14, 6–15, 6–16, 6–17, 6–18, 6–19, 6–20, 7–8, 7–9, 7–10, 7–11, 7–12, 7–13, 7–14, 7–15, 7–16, 7–17, 7–18, 7–19, 7–20, 8–9, 8–10, 8–11, 8–12, 8–13, 8–14, 8–15, 8–16, 8–17, 8–18, 8–19, 8–20, 9–10, 9–11, 9–12, 9–13, 9– 14, 9–15, 9–16, 9–17, 9–18, 9–19, 9–20, 10–11, 10–12, 10–13, 10–14, 10–15, 10–16, 10–17, 10–18, 10–19, 10–20, 11–12, 11–13, 11–14, 11–15, 11–16, 11–17, 11–18, 11–19, 11–20, 12–13, 310804994v3– 12 – 12–14, 12–15, 12–16, 12–17, 12–18, 12–19, 12–20, 13–14, 13–15, 13–16, 13–17, 13–18, 13–19, 13–20, 14–15, 14–16, 14–17, 14–18, 14–19, 14–20, 15–16, 15–17, 15–18, 15–19, 15–20, 16–17, 16–18, 16–19, 16–20, 17–18, 17–19, 17–20, 18–19, 18–20, 19–20) carbon atoms and, unless otherwise indicated, may be optionally substituted. In at least one embodiment, the alkyl is a 5 C1–C10 alkyl. In at least one embodiment, the alkyl is a C1–C6 alkyl. Suitable examples include, without limitation, methyl, ethyl, propyl, isopropyl, butyl, sec butyl, isobutyl, tert butyl, 3-pentyl, and the like.

[0079] As used herein, the term “alkenyl” refers to a straight or branched aliphatic unsaturated hydrocarbon of formula CnH2nhaving from two to about twenty (e.g., 2, 3, 4, 5, 6, 7,10 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 2–3, 2–4, 2–5, 2–6, 2–7, 2–8, 2–9, 2–10, 2–11, 2– 12, 2–13, 2–14, 2–15, 2–16, 2–17, 2–18, 2–19, 2–20, 3–4, 3–5, 3–6, 3–7, 3–8, 3–9, 3–10, 3–11, 3–12, 3–13, 3–14, 3–15, 3–16, 3–17, 3–18, 3–19, 3–20, 4–5, 4–6, 4–7, 4–8, 4–9, 4–10, 4–11, 4– 12, 4–13, 4–14, 4–15, 4–16, 4–17, 4–18, 4–19, 4–20, 5–6, 5–7, 5–8, 5–9, 5–10, 5–11, 5–12, 5– 13, 5–14, 5–15, 5–16, 5–17, 5–18, 5–19, 5–20, 6–7, 6–8, 6–9, 6–10, 6–11, 6–12, 6–13, 6–14, 6– 15 15, 6–16, 6–17, 6–18, 6–19, 6–20, 7–8, 7–9, 7–10, 7–11, 7–12, 7–13, 7–14, 7–15, 7–16, 7–17, 7–18, 7–19, 7–20, 8–9, 8–10, 8–11, 8–12, 8–13, 8–14, 8–15, 8–16, 8–17, 8–18, 8–19, 8–20, 9– 10, 9–11, 9–12, 9–13, 9–14, 9–15, 9–16, 9–17, 9–18, 9–19, 9–20, 10–11, 10–12, 10–13, 10–14, 10–15, 10–16, 10–17, 10–18, 10–19, 10–20, 11–12, 11–13, 11–14, 11–15, 11–16, 11–17, 11–18, 11–19, 11–20, 12–13, 12–14, 12–15, 12–16, 12–17, 12–18, 12–19, 12–20, 13–14, 13–15, 13–16, 20 13–17, 13–18, 13–19, 13–20, 14–15, 14–16, 14–17, 14–18, 14–19, 14–20, 15–16, 15–17, 15–18, 15–19, 15–20, 16–17, 16–18, 16–19, 16–20, 17–18, 17–19, 17–20, 18–19, 18–20, 19–20) carbon atoms in the chain and, unless otherwise indicated, may be optionally substituted. Exemplary alkenyls include, without limitation, ethylenyl, propylenyl, n-butylenyl, and i-butylenyl.

[0080] As used herein, the term “alkynyl” refers to a straight or branched aliphatic 25 unsaturated hydrocarbon of formula CnH2n-2having from two to about twenty (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 2–3, 2–4, 2–5, 2–6, 2–7, 2–8, 2–9, 2–10, 2–11, 2–12, 2–13, 2–14, 2–15, 2–16, 2–17, 2–18, 2–19, 2–20, 3–4, 3–5, 3–6, 3–7, 3–8, 3–9, 3–10, 3– 11, 3–12, 3–13, 3–14, 3–15, 3–16, 3–17, 3–18, 3–19, 3–20, 4–5, 4–6, 4–7, 4–8, 4–9, 4–10, 4–11, 4–12, 4–13, 4–14, 4–15, 4–16, 4–17, 4–18, 4–19, 4–20, 5–6, 5–7, 5–8, 5–9, 5–10, 5–11, 5–12, 30 5–13, 5–14, 5–15, 5–16, 5–17, 5–18, 5–19, 5–20, 6–7, 6–8, 6–9, 6–10, 6–11, 6–12, 6–13, 6–14, 6–15, 6–16, 6–17, 6–18, 6–19, 6–20, 7–8, 7–9, 7–10, 7–11, 7–12, 7–13, 7–14, 7–15, 7–16, 7–17, 7–18, 7–19, 7–20, 8–9, 8–10, 8–11, 8–12, 8–13, 8–14, 8–15, 8–16, 8–17, 8–18, 8–19, 8–20, 9– 10, 9–11, 9–12, 9–13, 9–14, 9–15, 9–16, 9–17, 9–18, 9–19, 9–20, 10–11, 10–12, 10–13, 10–14, 10–15, 10–16, 10–17, 10–18, 10–19, 10–20, 11–12, 11–13, 11–14, 11–15, 11–16, 11–17, 11–18, 310804994v3– 13 – 11–19, 11–20, 12–13, 12–14, 12–15, 12–16, 12–17, 12–18, 12–19, 12–20, 13–14, 13–15, 13–16, 13–17, 13–18, 13–19, 13–20, 14–15, 14–16, 14–17, 14–18, 14–19, 14–20, 15–16, 15–17, 15–18, 15–19, 15–20, 16–17, 16–18, 16–19, 16–20, 17–18, 17–19, 17–20, 18–19, 18–20, 19–20) carbon atoms in the chain and, unless otherwise indicated, may be optionally substituted. Exemplary 5 alkynyls include acetylenyl, propynyl, butynyl, 2-butynyl, 3-methylbutynyl, and pentynyl.

[0081] As used herein, the term “cycloalkyl” refers to a non-aromatic saturated or unsaturated monocyclic or polycyclic (e.g., bicyclyic, tricyclic, tetracyclic) ring system which may contain 3 to 24 (3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 3–4, 3–5, 3–6, 3–7, 3–8, 3–9, 3–10, 3–11, 3–12, 3–13, 3–14, 3–15, 3–16, 3–17, 3–18, 3–19, 3– 10 20, 3–21, 3–22, 3–23, 3–24, 4–5, 4–6, 4–7, 4–8, 4–9, 4–10, 4–11, 4–12, 4–13, 4–14, 4–15, 4–16, 4–17, 4–18, 4–19, 4–20, 4–21, 4–22, 4–23, 4–24, 5–6, 5–7, 5–8, 5–9, 5–10, 5–11, 5–12, 5–13, 5–14, 5–15, 5–16, 5–17, 5–18, 5–19, 5–20, 5–21, 5–22, 5–23, 5–24, 6–7, 6–8, 6–9, 6–10, 6–11, 6–12, 6–13, 6–14, 6–15, 6–16, 6–17, 6–18, 6–19, 6–20, 6–21, 6–22, 6–23, 6–24, 7–8, 7–9, 7–10, 7–11, 7–12, 7–13, 7–14, 7–15, 7–16, 7–17, 7–18, 7–19, 7–20, 7–21, 7–22, 7–23, 7–24, 8–9, 8– 15 10, 8–11, 8–12, 8–13, 8–14, 8–15, 8–16, 8–17, 8–18, 8–19, 8–20, 8–21, 8–22, 8–23, 8–24, 9–10, 9–11, 9–12, 9–13, 9–14, 9–15, 9–16, 9–17, 9–18, 9–19, 9–20, 9–21, 9–22, 9–23, 9–24, 10–11, 10–12, 10–13, 10–14, 10–15, 10–16, 10–17, 10–18, 10–19, 10–20, 10–21, 10–22, 10–23, 10–24, 11–12, 11–13, 11–14, 11–15, 11–16, 11–17, 11–18, 11–19, 11–20, 11–21, 11–22, 11–23, 11–24, 12–13, 12–14, 12–15, 12–16, 12–17, 12–18, 12–19, 12–20, 12–21, 12–22, 12–23, 12–24, 13–14, 20 13–15, 13–16, 13–17, 13–18, 13–19, 13–20, 13–21, 13–22, 13–23, 13–24, 14–15, 14–16, 14–17, 14–18, 14–19, 14–20, 14–21, 14–22, 14–23, 14–24, 15–16, 15–17, 15–18, 15–19, 15–20, 15–21, 15–22, 15–23, 15–24, 16–17, 16–18, 16–19, 16–20, 16–21, 16–22, 16–23, 16–24, 17–18, 17–19, 17–20, 17–21, 17–22, 17–23, 17–24, 18–19, 18–20, 18–21, 18–22, 18–23, 18–24, 19–20, 19–21, 19–22, 19–23, 19–24, 20–21, 20–22, 20–23, 20–24, 21–22, 22–23, 22–24, 23–24) carbon atoms, 25 which may include at least one double bond and, unless otherwise indicated, the ring system may be optionally substituted. Exemplary cycloalkyl groups include, without limitation, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, anti-bicyclopropane, and syn-bicyclopropane.

[0082] As used herein, the term “heterocycloalkyl” refers to a cycloalkyl group as defined 30 above having at least one O, S, and / or N interrupting the carbocyclic ring structure. Examples of heterocycloalkyls include, without limitation, piperidine, piperazine, morpholine, thiomorpholine, pyrrolidine, tetrahydrofuran, pyran, tetrahydropyran, and oxetane. Unless otherwise indicated, the heterocycloalkyl ring system may be optionally substituted. 310804994v3– 14 –

[0083] As used herein, the term “aryl” refers to an aromatic monocyclic or polycyclic (e.g., bicyclyic, tricyclic, tetracyclic) ring system from 6 to 24 (6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 6–7, 6–8, 6–9, 6–10, 6–11, 6–12, 6–13, 6–14, 6–15, 6–16, 6–17, 6–18, 6–19, 6–20, 6–21, 6–22, 6–23, 6–24, 7–8, 7–9, 7–10, 7–11, 7–12, 7–13, 7–14, 7–15, 7–16, 7–17, 5 7–18, 7–19, 7–20, 7–21, 7–22, 7–23, 7–24, 8–9, 8–10, 8–11, 8–12, 8–13, 8–14, 8–15, 8–16, 8– 17, 8–18, 8–19, 8–20, 8–21, 8–22, 8–23, 8–24, 9–10, 9–11, 9–12, 9–13, 9–14, 9–15, 9–16, 9–17, 9–18, 9–19, 9–20, 9–21, 9–22, 9–23, 9–24, 10–11, 10–12, 10–13, 10–14, 10–15, 10–16, 10–17, 10–18, 10–19, 10–20, 10–21, 10–22, 10–23, 10–24, 11–12, 11–13, 11–14, 11–15, 11–16, 11–17, 11–18, 11–19, 11–20, 11–21, 11–22, 11–23, 11–24, 12–13, 12–14, 12–15, 12–16, 12–17, 12–18, 10 12–19, 12–20, 12–21, 12–22, 12–23, 12–24, 13–14, 13–15, 13–16, 13–17, 13–18, 13–19, 13–20, 13–21, 13–22, 13–23, 13–24, 14–15, 14–16, 14–17, 14–18, 14–19, 14–20, 14–21, 14–22, 14–23, 14–24, 15–16, 15–17, 15–18, 15–19, 15–20, 15–21, 15–22, 15–23, 15–24, 16–17, 16–18, 16–19, 16–20, 16–21, 16–22, 16–23, 16–24, 17–18, 17–19, 17–20, 17–21, 17–22, 17–23, 17–24, 18–19, 18–20, 18–21, 18–22, 18–23, 18–24, 19–20, 19–21, 19–22, 19–23, 19–24, 20–21, 20–22, 20–23, 15 20–24, 21–22, 22–23, 22–24, 23–24) carbon atoms and, unless otherwise indicated, the ring system may be optionally substituted. Aryl groups of the present technology include, but are not limited to, groups such as phenyl, naphthyl, azulenyl, phenanthrenyl, anthracenyl, fluorenyl, pyrenyl, triphenylenyl, chrysenyl, naphthacenyl, biphenyl, triphenyl, and tetraphenyl. In at least one embodiment, an aryl within the context of the present technology is a 6 or 10 membered 20 ring. In at least one embodiment, each aryl is phenyl or naphthyl.

[0084] As used herein, the term “heteroaryl” refers to an aryl group as defined above having at least one O, S, and / or N interrupting the carbocyclic ring structure. Examples of heteroaryl groups include, without limitation, pyrrolyl, pyrazolyl, imidazolyl, triazolyl, furyl, thiophenyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, pyridyl, pyrazinyl, 25 pyrimidinyl, pyridazinyl, triazinyl, thienopyrrolyl, furopyrrolyl, indolyl, azaindolyl, isoindolyl, indolinyl, indolizinyl, indazolyl, benzimidazolyl, imidazopyridinyl, benzotriazolyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, pyrazolopyridinyl, triazolopyridinyl, thienopyridinyl, benzothiadiazolyl, benzofuyl, benzothiophenyl, quinolinyl, isoquinolinyl, tetrahydroquinolyl, tetrahydroisoquinolyl, cinnolinyl, quinazolinyl, quinolizilinyl, phthalazinyl, 30 benzotriazinyl, chromenyl, naphthyridinyl, acrydinyl, phenanzinyl, phenothiazinyl, phenoxazinyl, pteridinyl, and purinyl. Additional heteroaryls are described in Comprehensive Heterocyclic Chemistry: The Structure, Reactions, Synthesis and Use of Heterocyclic Compounds (Katritzky et al. eds., 1984), which is hereby incorporated by reference in its entirety. Unless otherwise indicated, the heteroaryl ring system may be optionally substituted. 310804994v3– 15 –

[0085] As used herein, the terms “alkoxy” refers to a groups of from 1 to 6 carbon atoms of a straight, branched, or cyclic configuration and combinations thereof attached to the parent structure through an oxygen. Examples include methoxy, ethoxy, propoxy, isopropoxy, butoxy, cyclopropyloxy, cyclohexyloxy, and the like. Alkoxy also includes methylenedioxy and 5 ethylenedioxy in which each oxygen atom is bonded to the atom, chain, or ring from which the methylenedioxy or ethylenedioxy group is pendant so as to form a ring. Thus, for example, phenyl substituted by alkoxy may be, for example,

[0086] As used herein, the term “aryloxy” refers to —OR, where R is an aryl group.

[0087] As used herein, the terms “perfluoroalkyl”, “perfluoroalkenyl”, “perfluoroalkynyl”, 10 and “perfluoroaryl” refer to an alkyl, alkenyl, alkynyl, or aryl group as defined above in which the hydrogen atoms on at least one of the carbon atoms have all been replaced with fluorine atoms.

[0088] The term “monocyclic” as used herein indicates a molecular structure having one ring. 15

[0089] The term “polycyclic” as used herein indicates a molecular structure having two or more rings, including, but not limited to, fused, bridged, spiro, or covalently bound rings. In at least one embodiment, the polycyclic ring system is a bicyclic, tricyclic, or tetracyclic ring system. In at least one embodiment, the polycyclic ring system is fused. In at least one embodiment, the polycyclic ring system is a bicyclic ring system such as naphthyl or biphenyl. 20

[0090] As used herein, the term “optionally substituted” indicates that a group may have a substituent at each substitutable atom of the group (including more than one substituent on a single atom), provided that the designated atom’s normal valency is not exceeded and the identity of each substituent is independent of the others. “Unsubstituted” atoms bear all of the hydrogen atoms dictated by their valency. When a substituent is keto (i.e., =O), then two 25 hydrogens on the atom are replaced. Combinations of substituents and / or variables are permissible only if such combinations result in stable compounds; by “stable compound” is meant a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture, and formulation into an efficacious agent.

[0091] As used herein, the term “unsubstituted” means that atoms bear all of the hydrogen 30 atoms dictated by their valency.

[0092] The sulfonamide probes of the present application can contain an aromatic or heteroaromatic chromophore. As used herein, an “aromatic or heteroaromatic chromophore” 310804994v3– 16 – refers to an aromatic or heteroaromatic group that produces a signal that can be used for chiroptical detection through various approaches including, without limitation, circular dichroism (CD), vibrational CD (VCD), electronic CD, optical rotatory dispersion (ORD), or polarimetry. 5

[0093] A UV chromophore shows a good absorption behavior in the spectral range of the UV rays or preferably an absorption maximum above 250nm. The chromophore absorbs the energy of the ultraviolet light and preferably does not change chemically as a result. The energy can be released as heat or phosphorescence / fluorescence. Visible chromophores include compounds with absorption from about 380nm to 740nm, which absorb light in the visible 10 spectrum. UV / Vis chromophores have a conjugated pi system, such as those found in aromatic compounds.

[0094] The analytical methods described herein may be used to evaluate chiral alcohols. The probes of the present application can covalently bond to the chiral alcohol in a reaction that replaces the alcohol moiety to form the probe-labeled analyte. In some embodiments, the probes 15 bond to the chiral alcohol via Mitsunobu reaction. In some embodiments, the chiral alcohol comprises a stereocenter at the site of the alcohol (–OH) moiety. In other embodiments, the chiral alcohol comprises an α-, β-, γ- or δ-stereocenter.

[0095] In some embodiments of the analytical methods described herein, the contacting step is carried out in a solvent selected from protic solvents, aprotic solvents, organic solvents, and 20 any combination thereof. Exemplary solvents include, but are not limited to, chloroform, dichloromethane, acetonitrile, toluene, tetrahydrofuran, methanol, ethanol, isopropanol, dimethyl sulfoxide (DMSO), dimethylformamide (DMF), pentane, pentane isomers, hexane, hexane isomers, ether, dichloroethane, acetone, ethyl acetate, butanone, and mixtures of any combination thereof. 25

[0096] In at least one embodiment of any analytical method described herein, contacting is carried out for about 30 to about 200 minutes (e.g., carried out for a duration range having an upper limit of about about 30, about 40, about 50, about 60, about 70, about 80, about 90, about 100, about 110, about 120, about 130, about 140, about 150, about 160, about 170, about 180, about 190, about 200, or any combination thereof). In all embodiments, contacting is carried out 30 for a time that is sufficient for the probe to bind to any analyte present in the sample. As will be apparent to the skilled chemist, the speed at which binding takes place will depend on various factors, including the particular probe selected and the analyte, whether a catalyst is present, concentrations, and the temperature. 310804994v3– 17 –

[0097] As will be apparent to the skilled chemist, the analytical methods may be carried out at room temperature, at high temperatures (e.g., about 50°C to about 100°C, e.g., a temperature range with an upper limit of about 55°C, about 60°C, about 65°C, about 70°C, about 75°C, about 80°C, about 85°C, about 90°C, about 95°C, or about 100°C, and a lower limit of about 50°C, 5 about 55°C, about 60°C, about 65°C, about 70°C, about 75°C, about 80°C, about 85°C, about 90°C, or about 95°C, or any combination thereof), or at low temperatures (e.g., below about 25°C, e.g., below about 25°C, below about 20°C, below about 15°C, below about 10°C, below about 5°C, below about 0°C, below about -5°C, below about -10°C, below about -15°C, below about -20°C, below about -25°C, below about -30°C, below about -35°C, below about -40°C,10 below about -45°C, below about -50°C, below about -55°C, below about -60°C, below about - 65°C, below about -70°C, or below about -75°C, preferably no lower than about -78°C; e.g., a temperature range with an upper limit of about 25°C, about 20°C, about 15°C, about 10°C, about 5°C, about 0°C, about -5°C, about -10°C, about -15°C, about -20°C, about -25°C, about -30°C, about -35°C, about -40°C, about -45°C, about -50°C, about -55°C, about -60°C, about -65°C, 15 about -70°C, or about -75°C, and a lower limit of about 20°C, about 15°C, about 10°C, about 5°C, about 0°C, about -5°C, about -10°C, about -15°C, about -20°C, about -25°C, about -30°C, about -35°C, about -40°C, about -45°C, about -50°C, about -55°C, about -60°C, about -65°C, about -70°C, about -75°C, or about -78°C, or any combination thereof). Furthermore, the analytical methods may be carried out under ambient conditions (e.g., 23±3° C. and 38±5% 20 relative humidity).

[0098] For example, the temperature could be increased to speed up the binding reaction. Some analyte–probe combinations may have side reactions at certain temperatures; the temperature could be decreased to prevent such side reactions.

[0099] In some embodiments, the contacting is carried out under a nitrogen atmosphere. 25

[0100] In some embodiments, the analytical methods are carried out in the presence of triphenylphosphine (PPh3) and diisopropyl azodicarboxylate (DIAD). In some embodiments, other azodicarboxylates can be used in please of DIAD. For example, in some embodiments, diethyl azodicarboxylate (DEAD), di-tert-butyl azodicarboxylate, or di-(4-chlorobenzyl) azodicarboxylate can be used in place of DIAD. 30

[0101] In the analytical methods described herein, the probe is reacted with the analyte to form probe-labeled analytes through a covalent bond between the probe and the analyte. The probe-labeled analytes generate a chiroptical signal that can be used to determine the concentration of the analyte in the sample and one or both of (i) the absolute configuration of the 310804994v3– 18 – analyte in the sample, and (ii) the enantiomeric and / or the diastereomeric composition of the analyte in the sample.

[0102] The chiroptical signal of the probe–analyte complexes can be measured using standard techniques, which will be apparent to the skilled artisan. Such techniques include 5 circular dichroism spectroscopy as well as the related vibrational circular dichroism and electronic circular dichroism spectroscopy formats (e.g., Stereochemistry of Organic Compounds 1003–07 (E. L. Eliel & S. H. Wilen eds., 1994); Dynamic Stereochemistry of Chiral Compounds 140–43 (Christian Wolf ed., 2008), each of which is hereby incorporated by reference in its entirety), optical rotatory dispersion (e.g., Stereochemistry of Organic Compounds 999–1003 (E. 10 L. Eliel & S. H. Wilen eds., 1994), which is hereby incorporated by reference in its entirety), and polarimetry (e.g., Stereochemistry of Organic Compounds 217–21, 1071–80 (E. L. Eliel & S. H. Wilen eds., 1994); Dynamic Stereochemistry of Chiral Compounds 140–43 (Christian Wolf ed., 2008), each of which is hereby incorporated by reference in its entirety). The absolute configuration of the analyte can also be assigned from the chiroptical signal of the probe–analyte 15 complexes that form. The configuration assignment can be based on the sense of chirality induction with a reference or by analogy.

[0103] The term “enantiomeric composition” refers to the enantiomeric ratio and / or enantiomeric excess of an analyte. The enantiomeric ratio (er) is the ratio of the percentage of one analyte enantiomer in a mixture to that of the other enantiomer. The enantiomeric excess 20 (ee) is the difference between the percentage of one analyte enantiomer and the percentage of the other analyte enantiomer. For example, a sample which contains 75% R- analyte and 25% S- analyte will have an enantiomeric excess of 50% of R- analyte and an enantiomeric ratio (S:R) of 25:75.

[0104] The term “diastereomeric composition” refers to the diastereomeric ratio and / or 25 diastereomeric excess of an analyte. The diastereomeric ratio is the ratio of the percentage of one analyte diastereomer in a mixture to that of the other diastereomer. The diastereomeric excess is the difference between the percentage of one analyte diastereomer and the percentage of the other analyte diastereomer. For example, a sample which contains 75% R,S-analyte and 25% S,S-analyte will have a diastereomeric excess of 50% of R,S-analyte and a diastereomeric 30 ratio (S,S:R,S) of 25:75.

[0105] In carrying out the analytical method, the step of contacting the sample with the probe to form a probe-labeled analyte is carried out in the presence of a known amount of excess triphenylphosphine, and determining the concentration of the chiral alcohol in the sample involves introducing a quinone compound into the reaction medium obtained following the 310804994v3– 19 – contacting step to cause a reaction between triphenylphosphine and the quinone compound that produces a colorimetric change. Quinones are a class of organic compounds which possess a fully conjugated cyclic dione structure. A suitable class of quinone probes is disclosed in U.S. Provisional Patent Application Serial No.63 / 173,071, which is hereby incorporated by reference 5 in its entirety. Exemplary quinones include, but are not limited to, 2-chloro-5-methyl-1,4- benzoquinone, 1,2-benzoquinone, 1,4-benzoquinone, 1,4-napthoquinone and 9,10- anthraquinone. An analog of a quinone is a quinone in which at least one of the hydrogen atoms has been replaced with a substituent including, but not limited to, a leaving group, a halogen, nitro, cyano, aryl, perfluoroaryl, heteroaryl, cycloalkyl, heterocycloalkyl, alkyl, or 10 perfluoroalkyl.

[0106] The colorimetric change is determined via UV-vis spectroscopy, wherein the colorimetric change is correlated to the amount of chiral alcohol in the sample. To determine the concentration of the analyte, the contacting step is carried out on at least three measurements to which different known concentrations of the analyte are introduced. One of the at least three 15 measurements comprises an excess concentration of the analyte, i.e., a saturating concentration that represents a maximum signal that can be detected in the chiroptical assay format used for detection. In certain embodiments, the at least three measurements can be carried out using four or more measurements, five or more measurements, six or more measurements, seven or more measurements, eight or more measurements, nine or more measurements, or ten or more 20 measurements. In general, the greater the number of measurements, then the more accurate the assessment of the concentration of the analyte in the sample.

[0107] In determining the concentration of the analyte in the sample, the intensity measurements obtained from the chiroptical assay (y-axis) are plotted against the analyte concentration (x-axis) for the at least three measurements to generate a calibration curve, and the 25 plotted data are then analyzed using a linear regression analysis.

[0108] In determining one or both of (i) the absolute configuration of the chiral alcohol in the sample, and (ii) the enantiomeric and / or the diastereomeric composition of the chiral alcohol in the sample, the step of contacting the sample with the probe to form a probe-labeled analyte is carried out on at least three measurements to which different known enantiomeric excesses of the 30 analyte are introduced. In certain embodiments, the at least three measurements can be carried out using four or more measurements, five or more measurements, six or more measurements, seven or more measurements, eight or more measurements, nine or more measurements, or ten or more measurements. In general, the greater the number of measurements, then the more accurate 310804994v3– 20 – the assessment of the enantiomeric and / or the diastereomeric composition of the analyte in the sample.

[0109] In determining the enantiomeric and / or the diastereomeric composition of the analyte in the sample, the intensity measurements obtained from the chiroptical assay (y-axis) are plotted 5 against the analyte enantiomeric excess (x-axis) for the at least three measurements to generate a calibration curve, and the plotted data are then analyzed using a linear regression analysis.

[0110] Preferences and options for a given aspect, feature, embodiment, or parameter of the technology described herein should, unless the context indicates otherwise, be regarded as having been disclosed in combination with any and all preferences and options for all other 10 aspects, features, embodiments, and parameters of the technology.

[0111] Another aspect of the present application relates to a kit for carrying out the analytical method described herein. The kit may include a solution comprising an aryl or heteroaryl sulfonamide probe as described herein and, optionally, one or more of (i) sample tubes suitable for use with a spectrophotometer; (ii) an optically pure reference sample of a chiral alcohol; (iii) 15 directions for using a spectrophotometer for carrying out circular dichroism (CD), vibrational CD (VCD), electronic CD, optical rotatory dispersion (ORD), or polarimetry analyses to measure the concentration of an analyte in a sample and one or both of the absolute configuration of the analyte in the sample, and the enantiomeric and / or the diastereomeric composition of the analyte in the sample; and (iv) a recordable medium comprising a template for analyzing data 20 obtained from the spectrophotometer and determining the concentration of an analyte in a sample and one or both of the absolute configuration of the analyte in the sample, and the enantiomeric and / or the diastereomeric composition of the analyte in the sample.

[0112] In addition to the foregoing, the kit may also include one or more, or all of the reagents used during the formation of the probe–analyte complexes and the reaction of the 25 reaction between triphenylphosphine and the quinone compound, as described herein. EXAMPLES

[0113] The examples below are intended to exemplify the practice of embodiments of the disclosure but are by no means intended to limit the scope thereof. 30 Example 1 – Screening of Probes 1-10 Against Chiral Alcohols

[0114] At the onset of this study, it was decided to screen the sulfonamides, imides and amides 1-10 equipped with extended chromophores and nitro groups or other auxochromes expected to generate strong CD effects when covalently attached to the chiral target molecules 310804994v3– 21 – via Mitsunobu reaction (FIGs.2A-2B, Materials and Methods). Note that all probes are achiral, an advantageous approach to avoid formation of diastereoisomers produced when chiral sensors are used which can complicate the stereochemical analysis and result in systematic errors. By contrast, the reaction of 1-10 with a chiral alcohol preserves the original enantiomeric ratio (er) 5 and CD spectroscopy can be used to determine er values directly from the intensity of the induced signals. CD spectroscopy is inherently primed to differentiate between molecular mirror images. Alcohols 11-22 were selected as test compounds to develop and evaluate the sensing assay. This group includes 2-butanol, 16, a particularly daunting sensing target that requires differentiation between a methyl and an ethyl group exhibiting very similar steric bulk at the 10 chiral center. Importantly, direct sensing of free alcohols 11-22 is not possible, because these compounds do not display quantifiable signals in the UV and CD regions of interest, which are generally accepted to be above 300 nm where interferences from impurities or reagents and by- products expected in multi-compound samples such as crude asymmetric reaction mixtures can be more easily excluded. 15

[0115] Several probes were found to fulfill this criterion and generate CD signals located at approximately 325 nm, including probes 1, 3, 4, and 6 (see FIG.7). Moreover, probe 3 gave strong CD inductions with all alcohols tested indicating unmatched utility to address the long- standing shortcomings in the chiroptical sensing realm listed in FIG.1. Reaction monitoring by 1H NMR spectroscopy and crystallographic analysis of an isolated product confirmed that the 20 alcohol transformation conducted with 3, PPh3and DIAD is quantitative and occurs with the expected configurational inversion (see Materials and Methods). The induced CD (ICD) effects obtained with the enantiomers of 1-phenylethanol, 11, and 2-octanol, 17, are shown exemplarily in FIG.2C (see Materials and Methods for chirality sensing of the other alcohols).

[0116] Probe 3 was then applied to samples containing both enantiomers of 2-butanol in 25 varying amounts, which revealed strong ICDs even with this challenging substrate and a linear relationship between the induced CD amplitude and the enantiomeric composition of 16 (FIG. 2D; see also Materials and Methods infra). The reaction is conveniently performed in acetonitrile / dichloromethane solution, but during CD optimization studies it was determined that the choice of the diluting solvent, which is added prior to the optical measurements, is important 30 (FIGs.2E and 10-14). While several solvents can be used, the strongest ICDs were generally observed with probe 3 in either THF, MeOH or CH2Cl2as shown in the corresponding heat map (FIG.2F). 310804994v3– 22 – Example 2 – Coupling CD Assay with UV Protocol to Allow Concomitant Optical er and Concentration Analysis

[0117] Having established a very sensitive chirality sensing protocol for alcohols, it was decided to couple the CD assay with a complementary UV protocol to enable concomitant 5 optical er and concentration analysis. In order to provide a user-friendly solution, the ability to integrate both tasks into a practical continuous workflow was sought. This is possible by exploiting PPh3in two ways (FIG.2B). When used in excess it can assist stoichiometrically in the Mitsunobu reaction and also serve as relay baton to connect the er sensing chemistry with UV concentration analysis based on previously reported redox chemistry with quinones 10 (Ramirez and Dershowitz, J. Am. Chem. Soc.78:5614–5622 (1956); Zhivetyeva et al., J. Fluor. Chem.180:21–32 (2015), which are hereby incorporated by reference in their entirety). This works very well with 27 yielding 28 and 29 as predicted by the literature and verified by ESI-MS analysis (see Materials and Methods and FIG.18). The chiroptical sensing relay thus produces a colorimetric change from red to yellow that is directly correlated to initial alcohol amount (FIG. 15 2G). Importantly, the UV indicator 27 can be added directly into the CD sensing solutions and samples are then processed continuously after 5 minutes without any additional treatment.

[0118] The performance of the relay assay was then evaluated with ten samples of 2-butanol covering a considerable range of total concentrations ([S]+[R]) and enantiomeric ratios ([S] / [R]) (Table 1). The chiroptical sensing method proved equally applicable to mixtures of small and 20 large er values. For example, the analysis of a 35.0 mM sample consisting of 62.0% (S)- and 38.0% (R)-16 gave 33.8 mM and an [S] / [R] ratio of 60.0:40.0 (entry 2). When a highly enantioenriched sample containing 8.0 mM of 2-butanol with an er of 98.0 (S):2.0 (R) was processed, the assay prediction was 7.5 mM with 98.5% (S)- and 1.5% (R)-16 (entry 7). Generally, accurate quantitative analysis was achieved with an averaged absolute error for the 25 concentration and enantiomeric composition determinations of + / -1.5 mM and 1.3%, respectively. In addition, the absolute configuration of the major enantiomer was correctly assigned in each case based on comparison of the sign of the ICD signal with a reference sample. 310804994v3– 23 – Table 1: Results of the chiroptical sensing of samples of 2-butanolAC=Absolute configuration, Conc.=Concentration. See Materials and Methods for details. Example 3 – Sensing of Challenging Alcohols Having β-, γ- or δ-Stereocenters

[0119] Encouraged by the strong ICD effects produced by 3 with the challenging alcohols 5 16-19 where both alkyl groups exhibit similar steric bulk at the chirality center, the possibility of chiroptical sensing of remote stereocenters was envisioned. Only one report discussing chirality sensing of γ-stereocenters has appeared in the literature (Dotson et al., J. Am. Chem. Soc. 143:19187-19198 (2021), which is hereby incorporated by reference in its entirety). In this work, a widely applicable Anslyn metal coordination assembly was refined using new components that 10 needed to be synthesized. While impressive proof-of-concept data were obtained, the assemblies consistently generated blue-shifted ICDs at low wavelengths below 300 nm, which limits the suitability for real-world samples that would contain UV- or CD-active interferents. To overcome these shortcomings and to extend the sensing scope further, 30-32 exhibiting a chirality center at the β-position were processed. Again, strong CD effects were generated in all 15 cases, prompting a more systematic investigation (FIGs.3A-3B).

[0120] For this purpose, the aliphatic scaffold present in 16 was evaluated. As mentioned above, stereodifferentiation between methyl and ethyl groups is a difficult task because of their very similar effective van der Waals radii, which is well documented by almost identical Taft and Charton steric parameters (Sigman and Miller, J. Org. Chem.74:7633-7643 (2009); Lin et 20 al., J. Am. Chem. Soc.138:8045-8047 (2016), which are hereby incorporated by reference in 310804994v3– 24 – their entirety). Compounds 33-35 systematically relocate the challenging sec-butyl unit in 16 into increasingly remote positions and display a growing degree of conformational flexibility, which is an additional obstacle that is generally detrimental to chiroptical sensing. Nevertheless, remote sensing of this motif in 33-35 is possible with this reaction-based chiroptical assay (FIGs.3A-3B 5 and Materials and Methods). Altogether, the relatively simple probe 3 has a truly unique application scope and allows chirality sensing of alcohols with α-, β-, γ- or even δ-stereocenters. Example 4 – Sensing of Challenging Alcohols Having β-, γ- or δ-Stereocenters

[0121] Enantiodifferentiation between deuterated isotopomers by small-molecule CD probes is another longstanding goal in the sensing field. Since Maeda, Yashima and coworkers showed 10 that this is possible through chiral amplification with 2,2’-biphenol-derived polyacetylene helices the utility of 3 was evaluated for this task (Maeda et al., J. Am. Chem. Soc.140:3270–3276 (2018), which is hereby incorporated by reference in its entirety). Following a modified literature protocol (Yamada and Noyori, Org. Lett.2:3425–3427 (2000), which is hereby incorporated by reference in its enitirety), the enantiomers of 36, which is chiral only by virtue of the H / D 15 isotopic substitution at the benzylic position, were prepared via Ru-catalyzed asymmetric reduction of the corresponding aldehyde with formic acid-d2(Matereials and Methods). In addition to the sensing of remote cryptochirality in 30-35 discussed above, this strategy to covalently place the optimized sulfonamide chromophore as close as possible or directly at the stereocenter as in 37 proves generally favorable for inducing quantifiable CD effects with these 20 challenging molecules. As shown in FIG.4, probe 3 is perfectly suited for highly sensitive isotopic enantiodifferentiation of 4-bromophenylmethan-d-ol. Chiral HPLC (Pirkle and Gan, Tetrahedron: Asymmetry 8:811–814 (1997); Kimata et al., J. Am. Chem. Soc.118:759–762 (1996), which are hereby incorporated by reference in their entirety), NMR (Meddour et al., J. Am. Chem. Soc.116:9652–9656 (1994); Lesot et al., Chem. Eur. J.10:3741-3746 (2004); 25 Naumann and Kuchel, Anal. Chem.87:10437–10442 (2015); Küppers et al., Eur. J. Org. Chem. 2019(15):2629–2634 (2019), which are hereby incorporated by reference in their enitirety) using chiral media, vibrational CD (Masarwa et al., Angew. Chem. Int. Ed.54:13106 –13109 (2015), which is hereby incorporated by reference in its entirety) and chiral tagging in molecular rotational resonance (MRR) (Domingos et al., Chem. Sci.11:10863–10870 (2020); Xie et al., 30 Angew. Chem., Int. Ed.59:15703–15710 (2020); Mills et al., Angew. Chem. Int. Ed. 61:e202207275 (2022); Vang et al., Chirality, 35:856–883 (2023); Sonstrom et al., Org. Process Res. Dev.27:1185–1197 (2023), which are hereby incorporated by reference in their entirety) spectroscopy are important techniques that have been applied to er and absolute configuration 310804994v3– 25 – analysis of molecules exhibiting enantio- or diastereomeric H / D substitution patterns. In particular, MRR spectroscopy which requires gas-phase measurements and quantum chemical analysis of the transition frequencies seems well-suited for deuterated isotopomer analysis. Despite impressive advances in the last few years, enantioselective MRR analysis takes close to 5 10 minutes per sample (Sonstrom et al., Org. Process Res. Dev.27:1185–1197 (2023), which is hereby incorporated by reference in its entirety) and is significantly more time-consuming than chiroptical sensing with automated CD plate readers which completes this task in only 3 seconds (Pilicer et al., J. Org. Chem.85:10858–10864 (2020), which is hereby incorporated by reference in its entirety). Moreover, MRR is limited to small molecules that are sufficiently volatile for the 10 gas-phase experimentation.

[0122] By contrast, the CD sensing method described herein is not restricted to small molecules like 11-22 or 30-36, but also inherently applicable to large compounds like cholesterol, 38, and the biologically active lignan podophyllotoxin, 39. See FIGs.39-40.

[0123] For comparison, the use of the well-known Mosher’s acid and a chiral lanthanide 15 shift reagent was investigated for enantioselective analysis of 33-36 (see Materials and Methods). It was observed that derivatization of alcohol 33 exhibiting a β-chirality center with Mosher’s acyl chloride forms diastereomeric esters that have different1H NMR signatures but are only partially resolved in the19F NMR spectrum. However, attempts to resolve the1H and 19F NMR signals of the diastereomeric esters formed with alcohols 34 and 35 having chirality 20 centers in the γ- and δ-positions, respectively, failed. Similarly, insufficient1H and lack of19F NMR signal resolution were observed when the Mosher esters of the deuterated alcohol 36 were investigated. Attempts to use the chiral lanthanide shift reagent Eu(facam)3also proved unsuccessful due to low peak resolution and signal broadening which is consistent with literature reports with chiral alcohols (Axt et al., J. Chem. Soc., Perkin Trans.2, 1999(12):2783–2788 25 (1999), which is hereby incorporated by reference in its entirety).

[0124] The ease of operation and unusual utility of this chiroptical relay sensing assay encouraged its employment without further adjustments in a real-world application with the goal to assess the robustness and general prospect of this methodology. Asymmetric reaction development generally requires screening of multiple parameters to identify optimal conditions 30 that give the desired product in high yields and enantiomeric excess. The need to evaluate potentially synergistic effects of different catalysts, solvents, additives etc. on the reaction outcome under steadily increasing time constraints that chemists nowadays have to adhere to make it highly desirable to investigate hundreds of parameter combinations in parallel to cover a broad chemical space literally overnight. In principal, this is possible with today’s high- 310804994v3– 26 – throughput experimentation equipment but the widespread use of traditional analytical techniques that are serial in nature and time-consuming remains a serious bottleneck. This can be addressed with chromatography-free chiroptical determination of product yield and enantiopurity as CD microplate readers have recently become commercially available. This would, of course, 5 necessitate chiroptical assays that are suitable for direct analysis of crude reaction mixtures. An autocatalytic asymmetric reaction (Chandrasekhar and Hota, Tetrahedron: Asymmetry 16:751– 754 (2005), which is hereby incorporated by reference in its entirety) that gives a chiral alcohol was therefore selected and a small portion of the product mixtures of three reactions run with different catalysts without any work-up and sample preparation was subjected to the UV / CD 10 assay (FIGs.5A-5B). Comparison with traditional GC methods showed that the conversion of 40 to 11 and the er of the chiral alcohol were determined with good accuracy and within an error range that is generally considered sufficient for high-throughput screening efforts aimed at rapid identification of optimal reaction conditions.

[0125] In summary, a practical reaction-based chiroptical sensing assay that allows 15 simultaneous determination of the concentration and enantiomeric purity of an unprecedented range of chiral alcohols has been developed. This is achieved with a conceptually new CD / UV relay sensing strategy that does not use the alcohol moiety for binding to a sensor, which is the generally followed practice, but instead replaces it with an optimized chromophoric sulfonamide unit to minimize the distance between the covalently incorporated chiroptical reporter and the 20 chirality center in the substrates. As demonstrated herein, this approach is conducive to strong CD and UV inductions at long wavelengths and enables enantiodifferentiation with very challenging targets exhibiting remote stereocenters or cryptochirality either by virtue of isotopic substitution or originating from sterically almost identical alkyl groups. The robustness and utility prospects of our sensing method were validated by chromatography-free asymmetric 25 reaction analysis with small aliquots of crude product mixtures. This replaces the common need for elaborate sample work-up and inherently slow one-sample-at-a-time handling protocols with a broadly useful optical method that offers increased speed, parallel sample processing, small- scale analysis capability and reduced chemical waste production. Based on the recent introduction of commercially available multi-well UV / CD plate readers, we expect that this 30 study paves the way toward high-throughput chiral compound development screening, a highly sought-after goal in the pharmaceutical industry where the synthesis of lead compounds often needs to be optimized by extensive reaction parameter optimization in a short time. 310804994v3– 27 – Materials and Methods for Examples 1-4

[0126] All commercially available reagents and solvents were used without further purification. Reactions and sensing studies were carried out under anhydrous conditions. Flash 5 chromatography was performed on silica gel, particle size 40-63 µm. NMR spectra were obtained on a Varian at 400 MHz (1H NMR) and 100 MHz (13C NMR) spectrometer at room temperature using either CDCl3 or CD3CN as solvent. Chemical shifts are reported in ppm relative to the solvent peaks. CD spectra were collected with a standard sensitivity of 100 mdeg, a data pitch of 1.0 nm, and a bandwidth of 1.0 nm in a continuous scanning mode with a 10 scanning speed of 200 nm / min and a response of 1.0 s (1 cm path length). The data were baseline corrected and smoothed using a binomial equation. GC-MS measurements were acquired on an Agilent 5977C GC / MSD equipped with an HP-5ms Ultra Inert (5% phenyl)methylpolysiloxane column (30 m, 0.25 mm, 0.25 µm). HR-MS data were obtained using electron spray ionization time-of-flight (ESI-TOF) spectrometry. 15 Probe Screening

[0127] Ten sulfonamides or amides carrying two aryl moieties were prepared as described hereinafter to identify a probe that would undergo quantitative Mitsunobu reaction and produce a strong red-shifted CD signal with an aliphatic alcohol substrate. For this purpose, a solution of (S)-1-methoxy-2-propanol (18.0 mg, 0.2 mmol) in 0.5 mL of THF was added to a solution of 20 PPh3(78.6 mg, 0.3 mmol) and DIAD (60.6 mg, 0.3 mmol) in 1.0 mL of THF under nitrogen (FIG.6). The mixture was stirred for 1 minute, followed by the addition of one of the amide or sulfonamide probes 1-10 (0.2 mmol) in 0.5 mL of CH2Cl2, bringing the total volume to 2.0 mL. After two hours of stirring the mixture was diluted to 2.0 mM with THF and subjected to CD analysis (FIG.7). NMR studies with 1 and 3 suggest that the reaction is complete within 1 hour 25 (FIGs.8 and 9). Probe 3 gave the strongest CD signal with a maximum of 100 mdeg at 325 nm at 2.0 mM in CH2Cl2. Thus, further optimization was carried out with 3. NMR Reaction Analysis

[0128] (S)-1-Phenylethanol (12.8 mg, 0.1 mmol) was added to a solution of PPh3(46.4 mg, 0.15 mmol) and DIAD (31.3 mg, 0.15 mmol) in 1.0 mL of CD3Cl under nitrogen. The mixture 30 was stirred for 1 minute, followed by the addition of 3 (49.5 mg, 0.15 mmol) in 0.5 mL of CD3Cl. The reaction was monitored by1H NMR spectroscopy and showed complete conversion within 1 hour (FIG.8). 310804994v3– 28 – Solvent Optimization

[0129] A solution of (S)-1-methoxy-2-propanol (9.0 mg, 0.1 mmol) in 0.5 mL of CH3CN was added to a vial containing PPh3(46.4 mg, 0.15 mmol) and DIAD (31.3 mg, 0.15 mmol) in 1.0 mL CH2Cl2:CH3CN (1:1, v / v) under nitrogen. The mixture was stirred for 1 minute, followed 5 by the addition of sulfonamide probe 3 (33.0 mg, 0.1 mmol) in 0.5 mL CH3CN, bringing the total volume to 2.0 mL. The mixture was stirred for 1 hour and then subjected to CD analysis at a concentration of 1.70 mM using CH3CN, CH2Cl2, DMSO, hexanes, methanol and THF as diluting solvent. Strong CD maxima at 325 mdeg were observed in all cases (FIG.10). Dilution with THF, MeOH and CH2Cl2produced the strongest CD signals. The use of DMSO and 10 CH3CN gave rise to weaker CD signals that would still suffice for quantitative ee analysis. Overall, THF gave the strongest CD signal and all subsequent CD spectra were collected using THF as diluting solvent. FIGs.11, 12, and 13 show the CD spectra of the sensing reaction of (S)- 1-methoxy-2-propanol with probes 1, 4, and 6, respectively. Confirmation of the Reaction Stereospecificity 15

[0130] To confirm that the reaction proceeds with stereoinversion, a single crystal of the reaction product from (R)-phenylethanol and N-(phenylsulfonyl)benzenesulfonamide was grown as described below. Single crystal analysis showed the formation of (S)-N-(1-phenylethyl)-N- (phenylsulfonyl)benzenesulfonamide with a Flack parameter of 0.004.

[0131] As shown if FIG.14, (R)-1-Phenylethanol (12.8 mg, 0.1 mmol) was added to a 20 solution of PPh3(46.43 mg, 0.15 mmol) and DIAD (31.3 mg, 0.15 mmol) in 1.0 mL of CHCl3:CH3CN (1:1, v / v) under nitrogen. The mixture was stirred for 1 minute, followed by the addition of 8 (29.7 mg, 0.1 mmol) in 0.5 mL of CH3CN. The mixture was allowed to stir for 1 hour at room temperature. The mixture was washed with water and extracted with CH2Cl2. The combined organic layers were dried over Na2SO4and concentrated in vacuo. Purification by 25 flash column chromatography on silica gel (25% ethyl acetate in hexanes) afforded 42.0 mg (0.1 mmol, 97%) of a white solid.

[0132] A single crystal was obtained by careful layering of pentane onto a solution of (S)-N- (1-phenylethyl)-N-(phenylsulfonyl)benzenesulfonamide in dichloromethane (pentane:CH2Cl2, 3:1, v / v). Single crystal analysis was performed at 100K using a Siemens platform diffractometer 30 with a graphite monochromated Mo-Kα radiation (λ = 0.71073 Å). Data were integrated and corrected using the APEX 3 program. The structures were solved by direct methods and refined with full-matrix least-square analysis using SHELXL-2019 / 1 software. Non-hydrogen atoms were refined with anisotropic displacement parameter. FIG.15 shows the crystal structure of (S)- N-(1-phenylethyl)-N-(phenylsulfonyl)benzenesulfonamide. Crystal data: C30N3O12S6, M= 310804994v3– 29 – 401.48, 0.292 x 0.176 x 0.152 mm3, orthorhombic, space group P212121, a= 13.3466(11), b= 16.19128 (14), c= 24.5648(19) Å, Z = 12. The CCDC number for the crystal reported in this study is 2288892. Analysis of the Phosphine Relay Reaction 5

[0133] The consumption of remaining PPh3upon completion of the Mitsunobu reaction serves as a relay indicator to determine the amount of the chiral alcohol (total concentration of both enantiomers). The unreacted PPh3which is used in excess in the chiroptical assay reacts rapidly with 27 to 28 and 29 thus producing a strong UV spectrum at 340 nm (Ramirez and Dershowitz, J. Am. Chem. Soc.78:5614–5622 (1956), which is hereby incorporated by reference 10 in its entirety). The more initial chiral alcohol is present in the sample, the less PPh3remains after reaction completion to react with 27. Using the following protocol, it was demonstrated that the change in the UV absorption correlates linearly with the initial chiral alcohol amount.

[0134] To a solution containing PPh3(60.0 mM) and DIAD (60.0 mM) in 300.0 µL of CH3CN:CH2Cl2(1:1, v / v), was added (S)-2-butanol in varying concentrations (0.0, 10.0, 20.0, 15 30.0 and 40.0 mM in CH3CN). The solution was allowed to stir for 1 minute, after which probe 3 (40.0 mM in CH3CN) was added, bringing the total volume to 500.0 µL. The reaction was complete after 1 hour. From each sample, a 100.0 µL aliquot was taken and added to a vial containing 40.0 µL of 27 (0.2 mM). The mixture was stirred for 5 minutes and subsequently subjected to UV analysis (1.5 µL of sample was diluted with 2.0 mL of CH3CN). The remaining 20 PPh3undergoes the reaction shown in FIG.16, producing a signal at 340 nm (FIG.17).

[0135] The outcome of the quinone / PPh3 relay reaction was verified by LC-MS. PPh3 (20.0 mg, 0.08 mmol) and 27 (47.8 mg, 0.3 mmol) were combined in 1.0 mL CH2Cl2. A 20.0 µL aliquot was diluted with 1.0 mL of CH3CN:H2O (3:1, v / v). Direct injection into a single quadrupole LC / MSD iQ showed two distinct species (FIG.18). The major species corresponds 25 to the reduced hydroquinone addition product 28. The reduction of 18 by PPh3is a competing pathway that leads to zwitterion 29’(Zhivetyeva et al., J. Fluor. Chem.180:21–32 (2015), which is hereby incorporated by reference in its entirety). We observed the protonated species 29 with the characteristic 3:1 M+2 chlorine isotope pattern. The UV spectra of the crude reaction mixture resemble previously reported spectra obtained from the reaction between PPh3and 2,5- 30 dichlorobenzoquinone (Ramirez and Dershowitz, J. Am. Chem. Soc.78:5614–5622 (1956), which is hereby incorporated by reference in its entirety). 310804994v3– 30 – Alcohol Substrate Scope

[0136] FIG.19 illustrates the reaction of probe 3 with alcohols 11-22, 30-36, 38, and 39. A solution of a chiral alcohol (0.02 mmol) in 0.1 mL of CH3CN was added to a vial containing PPh3(7.9 mg, 0.03 mmol) and DIAD (6.1 mg, 0.03 mmol) in 0.3 mL of CH2Cl2: CH3CN (1:1, 5 v / v) under nitrogen atmosphere. The mixture was stirred for 1 minute, followed by the addition of sulfonamide probe 3 (6.2 mg, 0.02 mmol) in 0.1 mL CH3CN, bringing the total volume to 0.5 mL. The mixture was stirred for 1 hour and then subjected to CD analysis (FIGs.20-40) after dilution with THF to the final concentration indicated under each figure (60.0-85.0 µL of the reaction mixture were added to 2.0 mL THF). 10 Quantitative Alcohol Sensing: Enantiomeric Excess and Total Concentration Sensing of 2-butanol

[0137] As previously described, the total concentration of 2-butanol can be determined by consumption of remaining PPh3with a quinone indicator. To a solution containing PPh3(60.0 15 mM) and DIAD (60.0 mM) in 300.0 µL CH3CN:CH2Cl2(1:1, v / v), was added (S)-2-butanol in varying concentrations (0.0, 10.0, 20.0, 30.0 and 40.0 mM in CH3CN). The solution was allowed to stir for 1 minute, after which probe 3 (40.0 mM in CH3CN) was added, bringing the total volume to 500.0 µL. The reaction was complete after 1 hour. From each sample, a 100.0 µL aliquot was taken and added to a vial containing 40.0 µL of 27 (0.2 M) in CH2Cl2. The mixture 20 was stirred for 5 minutes and subsequently subjected to UV analysis (1.5 µL of each sample was diluted in 2.0 mL of CH3CN) (FIG.41). A visible color gradient is observed from yellow to dark red depending on the amount of phosphine present (FIG.42). Plotting the intensity at 340 nm versus the concentration of 2-butanol yielded a straight line with R2=0.9901 and y=-0097x + 0.8211 (FIG.43). 25

[0138] The change in the CD amplitude upon addition of probe 3 to a solution of PPh3, DIAD, and 2-butanol was measured (FIG.44). Probe 3 (40.0 mM), phosphine (60.0 mM), DIAD (60.0 mM), and 2-butanol (total 40.0 mM) with varying ee’s (+100, +75, +50, +25, 0, -20, -50, - 75, and -100%) were dissolved in 500.0 µL of CH3CN:CH2Cl2(7:3, v / v) and stirred for 1 hour. Each mixture was diluted for CD analysis (75.0 µL aliquot diluted with 2.0 mL of THF). Plotting 30 the CD amplitude at 325 nm against the enantiomeric excess of 2-butanol yielded a straight line with R2=0.9996 and y=0.5077x + 1.3514 (FIG.45). Simultaneous determination of concentration and enantiomeric composition

[0139] Ten samples of 2-butanol at varying concentration and enantiomeric composition in CH2Cl2were prepared and subjected to simultaneous analysis of concentration, enantiomeric 310804994v3– 31 – excess and absolute configuration using probe 3. First, a UV spectrum was obtained as described above and the concentration was calculated using the intensities at 340 nm with the equation shown in FIG.43. Then, a CD spectrum was obtained as described above. The CD intensities were normalized to the concentration obtained from UV analysis and the ee was calculated using 5 the intensities at 325 nm and the equation shown in FIG.45. The absolute configuration was determined by comparing the sign of the Cotton effect to a reference sample. The concentrations, enantiomeric ratios, and absolute configurations determined for various samples of 2-butanol using this technique are summarized in Table 2. Table 2. Concentration, enantiomeric ratio, and absolute configuration of samples of 2- 10 butanol determined by simultaneous UV and CD responses of probe 3Analysis of asymmetric reductions of acetophenone

[0140] FIG.46 illustrates an autocatalytic reduction of acetophenone, and subsequent analysis via CD & UV spectroscopy and chiral GC-MS. The reaction was carried out using a 15 literature procedure (Chandrasekhar and Hota, Tetrahedron: Asymmetry 16:751–754 (2005), which is hereby incorporated by reference in its entirety). To a solution of NaBH4(9.1 mg, 0.24 mmol) in dry diglyme (1.0 mL) was added (-)-menthol (1.8 mg, 0.01 mmol) under nitrogen. The effervescent mixture was stirred for 20 minutes and 40 (24.1 mg, 0.2 mmol) was added. After 24 hours, the crude asymmetric reaction was subjected to chiroptical sensing and traditional GC-MS 20 protocols. The results of both chiroptical sensing and traditional GC-MS protocols are summarized in Table 3. 310804994v3– 32 – Table 3. Reaction analysis using either two GC-MS methods or chiroptical sensing. Catalyst Traditional Analysis Chiroptical Sensing Absolute Er (S:R) Conversion Absolute Er (S:R) Conversion Configuration Configuration C1 S 81.2:18.8 88.0 S 81.9:18.1 88.6 C2 S 65.5:34.5 87.3 S 67.9:32.1 90.2 C3 Rac 50.0:50.0 89.9 Rac 50.0:50.0 90.5 Chiroptical sensing

[0141] Upon completion of the ketone reduction reaction, a 100.0 µL aliquot was added to a 5 solution containing 400.0 µL of PPh3(60.0 mM), DIAD (60.0 mM) and probe 3 (40.0 mM). The mixture was stirred for 1 hour and subjected to UV and CD analysis as described above (FIGs. 47-50). Chiral GC-MS analysis

[0142] Upon completion, the reaction mixture was washed and extracted with CH2Cl2. The 10 combined organic layers were dried over Na2SO4and concentrated in vacuo. The desired compound was purified and isolated by flash column chromatography using hexanes-ethyl acetate (95:5) as mobile phase. The sample was diluted in CH2Cl2and subjected to both standard and chiral GC-MS analysis (FIGs.51-54). Conversion was determined by standard GC-MS, while enantiopurity was determined by chiral GC-MS. A chiral 2,6-methyl-3-pentyl-beta- 15 cyclodextrin (50% in polysiloxane) column was used with a constant oven temperature of 100 °C. (R)-Phenylethan-1-ol eluted at 16.9 min, while (S)-phenylethan-1-ol eluted at 19.1 min. Example 5 – Probe Synthesis and Characterization General Protocol for Sulfonamide and Benzamide Synthesis

[0143] To a solution of an amine (1.0 mmol), DIPEA (1.0 mmol) in 5.0 mL of CH2Cl2was 20 added a sulfonyl- or an acyl chloride (1.0 mmol). The mixture was allowed to stir for 1 hour at room temperature. Upon completion, the mixture was washed with water and extracted with CH2Cl2. The combined organic layers were dried over Na2SO4and concentrated in vacuo. The desired compound was purified and isolated by flash column chromatography using hexanes- ethyl acetate as mobile phase. 25

[0144] Probe characterization data for probes 1-7, 9, and 10 is presented below along with data for compounds 36 and 41-47. 310804994v3– 33 – N-Benzyl-2-nitrobenzenesulfonamide (1)

[0145] Probe 1 was obtained as an off-white oil in 95% yield (277.7 mg, 0.95 mmol) from benzylamine (107.1 mg, 1.0 mmol) and 2-nitrobenzenesulfonyl chloride (221.6 mg, 1.0 mmol) 5 following the general procedure described above using hexanes / EtOAc (80:20) as the mobile phase.1H NMR (400 MHz, CDCl3): δ= 7.98 (d, J = 7.6 Hz, 1H), 7.80 (d, J = 7.6 Hz, 1H), 7.69- 7.60 (m, 2H), 7.21 (m, 5H), 5.74 (t, J = 6.3 Hz, 1H), 4.32 (d, J = 6.3 Hz, 2H).13C NMR (100 MHz, CDCl3): δ= 147.8, 135.7, 133.9, 133.4, 132.7, 130.9, 128.6, 128.0, 127.8, 125.2, 47.8. The spectroscopic data are in accordance with the literature (Yu, et al., J. Org. Chem.87:14738– 10 14752 (2022), which is hereby incorporated by reference in its entirety). N-(2-Nitrobenzyl)-2-nitrobenzenesulfonamide (2)

[0146] Probe 2 was obtained as an off-white solid in 92% yield (310.3 mg, 0.92 mmol) from 15 2-nitrobenzylamine hydrochloride (188.61 mg, 1.0 mmol) and 2-nitrobenzenesulfonyl chloride (221.6 mg, 1.0 mmol) following the general procedure described above using hexanes / EtOAc (80:20) as the mobile phase.1H NMR (400 MHz, CDCl3): δ= 8.01-7.95 (m, 2H), 7.94 (d, J = 7.8 Hz, 1H), 7.71-7.59 (m, 4H), 7.44 (t, J = 7.8 Hz, 1H), 6.27 (t, J = 6.8 Hz, 1H), 4.62 (d, J = 6.8 Hz, 2H).13C NMR (100 MHz, CDCl3): δ= 147.9, 147.8, 134.1, 133.9, 133.6, 132.8, 132.0, 131.8, 20 130.6, 129.4, 125.5, 125.3, 45.6. HR-MS: (M+H)+, calc’d: 338.0369, found: 338.0437. 310804994v3– 34 – N-(4-Methoxyphenyl)-2-nitrobenzenesulfonamide (3)

[0147] Probe 3 was obtained as an orange solid in 92% yield (283.6 mg, 0.92 mmol) from p- anisidine (123.2 mg, 1.0 mmol) and 2-nitrobenzenesulfonyl chloride (221.6 mg, 1.0 mmol) 5 following the general procedure described above using hexanes / EtOAc (75:25) as the mobile phase.1H NMR (400 MHz, CD3CN): δ= 7.82 (d, J = 7.9 Hz, 1H), 7.73 – 7.66 (m, 2H), 7.55 (dd, J = 7.6 Hz, J = 7.6 Hz, 1H), 7.10 (bs, 1H), 7.06 (d, J = 8.8 Hz, 2H), 6.75 (d, J = 8.8 Hz, 2H), 3.73 (s, 3H).13C NMR (100 MHz, CDCl3): δ= 158.6, 148.2, 133.9, 132.5, 132.2, 131.9, 127.9, 126.2, 125.2, 114.6, 55.4. The spectroscopic data are in accordance with the literature (Yu, et al., 10 J. Org. Chem.87:14738–14752 (2022), which is hereby incorporated by reference in its entirety). N-(1-Naphthyl)-2-nitrobenzenesulfonamide (4)15

[0148] Probe 4 was obtained as a purple solid in 72% yield (283.6 mg, 0.92 mmol) from 1- aminonaphthalene (143.2 mg, 1.0 mmol) and 2-nitrobenzenesulfonyl (221.6 mg, 1.0 mmol) chloride following the general procedure described above using hexanes / EtOAc (75:25) as the mobile phase.1H NMR (400 MHz, CD3CN): δ= 8.05 (d, J = 7.7 Hz, 1H), 7.88 (d, J = 8.3 Hz, 1H), 7.82 (d, J = 7.7 Hz, 1H), 7.77 (d, J = 7.7 Hz, 1H), 7.66 (m, 2H), 7.57 (bs, 1H), 7.51 – 7.33 20 (m, 5H).13C NMR (100 MHz, CDCl3): δ= 148.1, 134.3, 133.8, 133.0, 132.6, 131.4, 130.9, 129.7, 128.4, 128.2, 126.9, 126.5, 125.3, 125.2, 124.2, 121.8. The spectroscopic data are in accordance with the literature (Sokolov et al., Russ. J. Org. Chem.41:1023–1035 (2005), which is hereby incorporated by reference in its entirety). 310804994v3– 35 – N-(4-Methoxyphenyl)-3-nitrobenzesulfonamide (5)

[0149] Probe 5 was obtained as a white solid in 98% yield (266.8 mg, 0.98 mmol) from p- anisidine (123.2 mg, 1.0 mmol) and 3-nitrobenzenesulfonyl chloride (221.6 mg, 1.0 mmol) 5 following the general procedure described above using hexanes / EtOAc (85:15) as the mobile phase.1H NMR (400 MHz, CDCl3): δ = 8.56 (dd, J = 2.0 Hz, J = 2.0 Hz, 1H), 8.37 (m, 1H), 7.95 (m, 1H), 7.62 (dd, J = 8.1 Hz, J = 8.1 Hz, 1H), 7.01 – 6.93 (m, 2H), 6.80 – 6.74 (m, 2H), 3.75 (s, 3H).13C NMR (100 MHz, CDCl3): δ= 158.6, 148.2, 141.2, 132.8, 130.2, 127.5, 127.3, 126.0, 122.5, 114.7, 55.4. The spectroscopic data are in accordance with the literature (Eufrásio et al., 10 ACS Infect. Dis.7:2455–2471 (2021), which is hereby incorporated by reference in its entirety). N-(4-Methoxyphenyl)-2.4-dinitrobenzenesulfonamide (6)

[0150] Probe 6 was obtained as an orange solid in 94% yield (331.8 mg, 0.94 mmol) from p- 15 anisidine (123.2 mg, 1.0 mmol) and 2,4-dinitrobenzenesulfonyl chloride (266.6 mg, 1.0 mmol) following the general procedure described above using hexanes / EtOAc (75:25) as the mobile phase.1H NMR (400 MHz, CDCl3): δ= 8.66 (m, 1H), 8.37 (m, 1H), 8.01 – 7.92 (m, 1H), 7.16 (bs, 1H), 7.13 – 7.05 (m, 2H), 6.85 – 6.76 (m, 2H), 3.77 (s, 3H).13C NMR (100 MHz, CDCl3): δ= 161.7, 152.7, 151.1, 140.5, 136.3, 129.5, 129.3, 129.0, 123.2, 117.5, 59.2. The spectroscopic 20 data are in accordance with the literature (Malwal et al., J. Med. Chem.55:553–557 (2012), which is hereby incorporated by reference in its entirety). 310804994v3– 36 – N-(4-Methoxyphenyl)-4-methylbenzenesulfonamide (7)

[0151] Probe 7 was obtained as a white solid in 96% yield (266.2 mg, 0.96 mmol) from p- anisidine (123.2 mg, 1.0 mmol) and p-toluenesulfonyl chloride (190.7 mg, 1.0 mmol) following 5 the general procedure described above using hexanes / EtOAc (85:15) as the mobile phase.1H NMR (400 MHz, CDCl3): δ= 7.62 – 7.55 (d, J = 7.9 Hz, 2H), 7.20 (d, J = 7.9 Hz, 2H), 7.01 – 6.92 (d, J = 7.8 Hz, 2H), 6.79 – 6.70 (d, J = 7.8 Hz, 2H), 6.63 (bs, 1H), 3.74 (s, 3H), 2.37 (s, 3H). 13C NMR (100 MHz, CDCl3): δ= 157.9, 143.7, 135.9, 129.5, 128.9, 127.3, 125.4, 114.4, 55.4, 21.5. The spectroscopic data are in accordance with the literature (Lei et al., Org. Lett.25:6001– 10 6005 (2023), which is hereby incorporated by reference in its entirety). N-(4-Methoxyphenyl)-2-nitrobenzamide (9)

[0152] Probe 9 was obtained as a white solid in 98% yield (266.8 mg, 0.98 mmol) from p- 15 anisidine (123.2 mg, 1.0 mmol) and 2-nitrobenzoyl chloride (221.6 mg, 1.0 mmol) following the general procedure described above using hexanes / EtOAc (85:15) as the mobile phase.1H NMR (400 MHz, CDCl3): δ= 8.12 (d, J = 8.4 Hz, 1H), 7.73 (m, 1H), 7.65-7.60 (m, 2H), 7.50 (d, J = 8.6 Hz, 2H), 7.36 (bs, 1H), 6.95 – 6.88 (m, 2H), 3.82 (s, 3H).13C NMR (100 MHz, CDCl3): δ= 164.3, 157.2, 133.9, 132.9, 130.7, 130.2, 129.7, 128.7, 124.7, 122.5, 114.3, 55.5. The 20 spectroscopic data are in accordance with the literature (Bao et al., Org. Lett.22:6277–6282 (2020), which is hereby incorporated by reference in its entirety). 310804994v3– 37 – (E)-N-(4-((4-dimethylamino)phenyl)diazenyl)phenyl)-2-nitrobenzenesulfonamide (10)

[0153] Probe 10 was obtained as a red solid in 45% yield (191.5 mg, 0.45 mmol) from 4- amino-4’-dimethylaminoazobenzene (240.3 mg, 1.0 mmol) and 2-nitrobenzenesulfonyl (221.6 5 mg, 1.0 mmol) chloride following the general procedure described above using hexanes / EtOAc (75:25) as the mobile phase.1H NMR (400 MHz, CDCl3): δ= 7.86-7.84 (m, 4H), 7.77-7.75 (m, 2 H), 7.67 (dd, J = 7.7 Hz, J = 7.7 Hz, 1H), 7.56 (dd, J = 7.7 Hz, J = 7.7 Hz, 1H), 7.34 (bs, 1H), 7.31-7.29 (m, 2H), 6.77 (d, J = 8.8 Hz, 2H), 3.10 (s, 6H).13C NMR (100 MHz, CDCl3): δ= 152.9, 148.2, 136.1, 134.0, 132.6, 132.1, 131.9, 125.3, 123.9, 123.4, 123.2, 122.9, 111.7, 40.4. 10 HR-MS: (M+H)+, calc’d: 426.1236, found: 426.1256. Example 6 – Synthesis and Characterization of Compounds 36 and 41-47 N-(4-Methoxyphenyl)-N-(1-phenylethyl)-2-nitrobenzenesulfonamide (41)

[0154] (S)-1-Phenylethanol (12.8 mg, 0.11 mmol) was added to a solution of PPh3(46.43 15 mg, 0.16 mmol) and DIAD (31.3 µl, 0.16 mmol) in 1 mL of CHCl3: CH3CN (1:1, v / v) under nitrogen. The mixture was stirred for 1 minute, followed by the addition of probe 3 (33.0 mg, 0.11 mmol) in 0.5 mL of CH3CN. The mixture was allowed to stir for 1 hour at room temperature. The mixture was washed with water and extracted with CH2Cl2. The combined organic layers were dried over Na2SO4and concentrated in vacuo. Purification by flash column 20 chromatography on silica gel (25% ethyl acetate in hexanes) afforded 42.0 mg (0.1 mmol, 97%) of an orange oil.1H NMR (400 MHz, CDCl3): δ= 7.67 – 7.59 (m, 2H), 7.53 (m, 1H), 7.47 (m, 1H), 7.26 – 7.23 (m, 5H), 6.67 (d, J = 8.5 Hz, 2H), 6.60 (d, J = 8.5 Hz, 2H), 5.77 (q, J = 7.2 Hz, 1H), 3.75 (s, 3H), 1.54 (d, J = 7.0 Hz, 3H).13C NMR (100 MHz, CDCl3): δ= 159.4, 147.8, 140.2, 310804994v3– 38 – 134.3, 133.3, 132.1, 131.1, 128.3, 128.2, 127.9, 126.1, 123.9, 113.7, 58.0, 55.4, 18.5. HR-MS: (M+H)+, calc’d: 412.1093, found: 412.1072. Asymmetric synthesis of the enantiomers of 4-bromophenylmethan-d-ol (36) 5

[0155] The reaction was carried out using a modified literature procedure (Yamada and Noyori, Org. Lett.2:3425–3427 (2000), which is hereby incorporated by reference in its entirety). The Ru catalyst (6.4 mg, 0.01 mmol), formic acid-d2(101.1 mg, 2.0 mmol), and 10 triethylamine (202.4 mg, 2.0 mmol) were dissolved in 1.0 mL of anhydrous acetonitrile under nitrogen. The mixture was stirred for 10 minutes followed by the addition of 4- bromobenzaldehyde (370.0 mg, 2.0 mmol). Upon reaction completion after 18 hours, the mixture was washed with water and brine, and extracted with ethyl acetate. The combined organic layers were dried over Na2SO4and concentrated in vacuo. Purification by flash column 15 chromatography on silica gel (5% ethyl acetate in hexanes) afforded 357.2 mg (1.9 mmol, 95%) of a white solid. The ee was determined as >99% by chiral GC-MS using a chiral 2,6-dimethyl- 3-pentyl-β-cyclodextrin (50% in polysiloxane) capillary with a constant oven temperature of 130 °C (FIGs.55-57).1H NMR (400 MHz, CDCl3) δ = 7.47 – 7.39 (d, J = 8.3 Hz, 2H), 7.19 – 7.12 (d, J = 8.3 Hz, 2H), 4.54 (s, 1H), 2.46 (bs, 1H).13C NMR (100 MHz, CDCl3) δ = 139.66, 131.57, 20 128.58, 121.40, 64.04 (t, JC-D= 21.8 Hz). The spectroscopic data are in accordance with the literature (Yamada and Noyori, Org. Lett.2:3425–3427 (2000), which is hereby incorporated by reference in its entirety). The other enantiomer was prepared following the same protocol with the enantiomeric Ru catalyst. 25 Analysis of cryptochirality with Mosher’s acyl chloride and Eu(facam)3

[0156] The reaction was carried out using a modified literature procedure (Dale et al., J. Org. Chem.9:2543-2549 (1969), which is hereby incorporated by reference in its entirety). To a solution of (S) -Mosher’s acyl chloride (20mg, 0.07 mmol) in 1.0 mL CH2Cl2was added chiral alcohol (0.07 mmol) and DIPEA (9.62, 0.07 mmol). The mixture was stirred for 1.5 hours. Upon 310804994v3– 39 – reaction completion, the mixture was washed with water and brine, and extracted with ethyl acetate. The combined organic layers were dried over Na2SO4and concentrated in vacuo. Purification by flash chromatography on silica gel (5% ethyl acetate in hexanes) afforded the desired product. 5 2-Methylbutyl (2S)-2-methoxy-2-phenyl-2-(trifluoromethoxy)acetate (42)

[0157] Compound 42 was obtained as a colorless oil in 95% yield (22.7 mg, 0.07 mmol) 10 from (S)-2-methoxy-2-phenyl-2-(trifluoromethoxy)acetyl chloride (20.0 mg, 0.07 mmol) and racemic 2-methylbutan-1-ol (6.6 mg, 0.07 mmol) following the general procedure described above.1H NMR (400 MHz, CDCl3): δ= 7.52-7.48 (m, 2H), 7.43-7.33 (m, 3H), 4.23 (dd, J= 10.7, 5.7 Hz, 0.5 H), 4.15 (d, J= 6.1 Hz, 1H), 4.07 (dd, J= 10.7, 6.6 Hz, 0.5 H), 3.54 (s, 3H), 1.77 (m, 1H), 1.38 (m, 1H), 1.17 (m, 1H), 0.91-0.85 (m, 6H).19F NMR (376 MHz, CDCl3): δ= -71.63 (d, 15 3F). (S)-2-Methylbutyl (2S)-2-methoxy-2-phenyl-2-(trifluoromethoxy)acetate ((S,S)-42)(S)-3320

[0158] (S,S)-42 was obtained as a colorless oil in 94% yield (22.5 mg, 0.07 mmol) from (S)- 2-methoxy-2-phenyl-2-(trifluoromethoxy)acetyl chloride (20.0 mg, 0.07 mmol) and (S)-2- methylbutan-1-ol (6.6 mg, 0.07 mmol) following the general procedure described above.1H NMR (400 MHz, CDCl3): δ= 7.49-7.47 (m, 2H), 7.43-7.33 (m, 3H), 4.23 (dd, J= 10.7, 5.7 Hz, 1 H), 4.07 (dd, J= 10.7, 6.6 Hz, 0.5 H), 3.54 (s, 3H), 1.71 (m, 1H), 1.39 (m, 1H), 1.18 (m, 1H), 25 0.93-0.84 (m, 6H).19F NMR (376 MHz, CDCl3): δ= -71.62 (s, 3F). 3-Methylpentyl (2S)-2-methoxy-2-phenyl-2-(trifluoromethoxy)acetate (43)310804994v3– 40 –

[0159] Compound 43 was obtained as a colorless oil in 94% yield (23.3 mg, 0.07 mmol) from (S)-2-methoxy-2-phenyl-2-(trifluoromethoxy)acetyl chloride (20.0 mg, 0.07 mmol) and racemic 3-methylpentan-1-ol (7.6 mg, 0.07 mmol) following the general procedure described above.1H NMR (400 MHz, CDCl3): δ= 7.54-7.47 (m, 2H), 7.43-7.33 (m, 3H), 4.42-4.25 (m, 2 5 H), 3.54 (s, 3H), 1.71 (m, 1H), 1.59-1.23 (m, 3H), 1.15 (m, 1H), 0.90-0.81 (m, 6H).19F NMR (376 MHz, CDCl3): δ= -71.65 (s, 3F). (S)-3-Methylpentyl (S)-2-methoxy-2-phenyl-2-(trifluoromethoxy)acetate ((S,S)-43)10

[0160] Compound (S,S)-43) was obtained as a colorless oil in 94% yield (23.2 mg, 0.07 mmol) from (S)-2-methoxy-2-phenyl-22-(trifluoromethoxy)acetyl chloride (20.0 mg, 0.07 mmol) and (S)-3-methylpentan-1-ol (7.6 mg, 0.07 mmol) following the general procedure described above.1H NMR (400 MHz, CDCl3): δ= 7.52-7.49 (m, 2H), 7.40-7.35 (m, 3H), 4.42-4.25 (m, 2 15 H), 3.54 (s, 3H), 1.71 (m, 1H), 1.55-1.24 (m, 3H), 1.15 (m, 1H), 0.90-0.79 (m, 6H).19F NMR (376 MHz, CDCl3): δ= -71.65 (s, 3F). 4-Methylhexyl (S)-2-methoxy-2-phenyl-2-(trifluoromethoxy)acetate (44)20

[0161] Compound 44 was obtained as a colorless oil in 96% yield (24.8 mg, 0.07 mmol) from (S)-2-methoxy-2-phenyl-22-(trifluoromethoxy)acetyl chloride (20.0 mg, 0.07 mmol) and racemic 4-methylhexan-1-ol (8.6 mg, 0.07 mmol) following the general procedure described above.1H NMR (400 MHz, CDCl3): δ= 7.54-7.45 (m, 2H), 7.40-7.38 (m, 3H), 4.35-4.23 (m, 2 25 H), 3.54 (s, 3H), 1.75-1.58 (m, 2H), 1.38-1.22 (m, 3H), 1.15-1.06 (m, 2H), 0.84-0.80 (m, 6H). 19F NMR (376 MHz, CDCl3): δ= -71.65 (s, 3F). (S)-4-Methylhexyl (S)-2-methoxy-2-phenyl-2-(trifluoromethoxy)acetate ((S,S)-44)310804994v3– 41 –

[0162] Compound (S,S)-44 was obtained as a colorless oil in 97% yield (25.0 mg, 0.07 mmol) from (S)-2-methoxy-2-phenyl-22-(trifluoromethoxy)acetyl chloride (20.0 mg, 0.07 mmol) and (S)-4-methylhexan-1-ol (8.6 mg, 0.07 mmol) following the general procedure described above.1H NMR (400 MHz, CDCl3): δ= 7.54-7.48 (m, 2H), 7.40-7.38 (m, 3H), 4.35-4.23 (m, 2 5 H), 3.54 (s, 3H), 1.75-1.58 (m, 2H), 1.38-1.22 (m, 3H), 1.15-1.06 (m, 2H), 0.84-0.80 (m, 6H). 19F NMR (376 MHz, CDCl3): δ= -71.65 (s, 3F). (4-Bromophenyl)methyl-d (2S)-2-methoxy-2-phenyl-2-(trifluoromethoxy)acetate (45)10

[0163] Compound 50 was obtained as a colorless oil in 94% yield (27.6 mg, 0.066 mmol) from (S)-2-methoxy-2-phenyl-22-(trifluoromethoxy)acetyl chloride (20.0 mg, 0.07 mmol) and racemic (4-bromophenyl)methan-d-ol (13.2 mg, 0.07 mmol) following the general procedure described above.1H NMR (400 MHz, CDCl3): δ= 7.50-7.30 (m, 7H), 7.23-7.16 (m, 2H), 5.25 (d, 15 J= 5.9 Hz, 1H), 3.49 (s, 3H).19F NMR (376 MHz, CDCl3): δ= -71.68 (s, 3F). NMR analysis with Eu(facam)320

[0164] The reaction was carried out using a modified literature procedure (Goering et al., J. Am. Chem. Soc.5:1493–1501 (1974), which is hereby incorporated by reference in its entirety). To a solution of 46 (10.0 mg, 0.01 mmol) in 1.0 mL CDCl3was added racemic 34 (0.01 mmol). The mixture was stirred for 30 minutes.1H NMR analysis showed broad unresolved peaks which is consistent with literature reports for chiral alcohols (Axt et al., J. Chem. Soc., Perkin Trans.2, 25 1999(12):2783–2788 (1999), which is hereby incorporated by reference in its entirety).

[0165] Although preferred embodiments have been depicted and described in detail herein, it will be apparent to those skilled in the relevant art that various modifications, additions, substitutions, and the like can be made without departing from the spirit of the invention and 310804994v3– 42 – these are therefore considered to be within the scope of the invention as defined in the claims which follow. 5 310804994v3

Claims

– 43 – WHAT IS CLAIMED IS:

1. An analytical method comprising: providing a sample potentially containing a chiral alcohol that can exist in stereoisomeric forms; 5 providing an aryl or heteroaryl sulfonamide probe, contacting the sample with the probe, wherein said contacting is carried out under conditions to permit reaction of the probe with the chiral alcohol, if present in the sample, to form a probe-labeled analyte; and detecting the probe-labeled analyte in the sample using one or more chiroptical assay 10 formats, and determining the concentration of the chiral alcohol in the sample and one or both of (i) the absolute configuration of the chiral alcohol in the sample, and (ii) the enantiomeric and / or the diastereomeric composition of the chiral alcohol in the sample.

2. The analytical method according to claim 1, wherein the probe covalently bonds to the 15 chiral alcohol in a reaction that replaces the alcohol moiety to form the probe-labeled analyte.

3. The analytical method according to claim 1 or 2, wherein the probe is an achiral sulfonamide compound having the structure according to formula (I):20 wherein Ar is a substituted or unsubstituted aromatic or heteroaromatic group, R1is H, a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl, R2is a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl, and n is 0, 1, 2, 3, 4, or 5. 25 4. The analytical method according to claim 3, wherein the substituted or unsubstituted aryl at Ar, R1, and R2is independently selected from the group consisting of phenyl, naphthyl, azulenyl, phenanthrenyl, anthracenyl, fluorenyl, pyrenyl, triphenylenyl, chrysenyl, naphthacenyl, biphenyl, triphenyl, and tetraphenyl. 30 5. The analytical method according to claim 3, wherein the substituted or unsubstituted heteroaryl at Ar, R1, and R2is independently selected from the group consisting of pyrrolyl, pyrazolyl, imidazolyl, triazolyl, furyl, thiophenyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, triazinyl, thienopyrrolyl, 310804994v3– 44 – furopyrrolyl, indolyl, azaindolyl, isoindolyl, indolinyl, indolizinyl, indazolyl, benzimidazolyl, imidazopyridinyl, benzotriazolyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, pyrazolopyridinyl, triazolopyridinyl, thienopyridinyl, benzothiadiazolyl, benzofuyl, benzothiophenyl, quinolinyl, isoquinolinyl, tetrahydroquinolyl, tetrahydroisoquinolyl, cinnolinyl, 5 quinazolinyl, quinolizilinyl, phthalazinyl, benzotriazinyl, chromenyl, naphthyridinyl, acrydinyl, phenanzinyl, phenothiazinyl, phenoxazinyl, pteridinyl, and purinyl.

6. The analytical method according to claim 3, wherein Ar is selected from:10 wherein: the squiggly line represents the point of attachment to the −S(O2)− group in formula I; each X is independently C or N, except that no more than three ring nitrogens are present in the substituted or unsubstituted aromatic or heteroaromatic ring, and R3, R4, R5, R6, and R7are independently selected from the group consisting of a lone pair (when 15 X is N), –H, –CN, –NO2, halogen, –C1-C6alkyl, –C1-C6alkoxy, –N–(alkyl)2, –C1-C6alkenyl, –C1-C6alkynyl, – C1-C6perfluoroalkyl, –aryl, –perfluoroaryl, –aryloxy, –N–(aryl)2, –heteroaryl, –O–heteroaryl, –N–(heteroaryl)2, –cycloalkyl, –O–cycloalkyl, –N–(cycloalkyl)2, –heterocycloalkyl, –O–heterocycloalkyl, –N–(heterocycloalkyl)2, –C(O)Ra, –SO2Ra, and –OC(O)Ra; 20 each Ra is independently selected from the group consisting of –alkyl, –O–alkyl, –N–(alkyl)2, –alkenyl, –alkynyl, –aryl, –O–aryl, –N–(aryl)2, –heteroaryl, –O–heteroaryl, –N–(heteroaryl)2, –cycloalkyl, –O–cycloalkyl, –N–(cycloalkyl)2, –heterocycloalkyl, –O–heterocycloalkyl, and –N–(heterocycloalkyl)2; and wherein, optionally, R3and R4, R4and R5, R5and R6, and / or R6and R7are alternatively taken 25 together with the carbon or nitrogen atoms to which they are attached to form a fused monocyclic or bicyclic ring system selected from the group consisting of cycloalkyl, heterocycloalkyl, aryl, and heteroaryl, wherein the fused ring system is optionally substituted with one or more groups selected from –alkyl, –O–alkyl, –N–(alkyl)2, – alkenyl, –alkynyl, –O-aryl, –O-heteroaryl, –N-(aryl)2, –N-(heteroaryl)2, –aryl, –C(O)Rc, 310804994v3– 45 – –CO2Rb, –O-C(O)Rb, –NHC(O)Rb, –NRcC(O)Rb, –NO2, –CN, –halogen, and –SO2Rb, wherein each Rbis independently aryl, heteroaryl, alkyl, or CH2Ar.

7. The analytical method according to any one of claims 1–6, wherein the probe is selected 5 from:

8. The analytical method according to any one of claims 1–7, wherein said contacting the 10 sample with the probe is carried out via Mitsunobu reaction.

9. The analytical method according to any one of claims 1–8, wherein the chiral alcohol comprises a stereocenter at the site of the alcohol (–OH) moiety. 15 10. The analytical method according to any one of claims 1–9, wherein the stereocenter is an α-, β-, γ- or δ-stereocenter.

11. The analytical method according to any one of claims 1–10, wherein the chiroptical assay format is circular dichroism (CD), vibrational CD (VCD), electronic CD, optical rotatory 20 dispersion (ORD), or polarimetry.

12. The analytical method according to any one of claims 1–11, wherein said contacting is carried out in a solvent selected from protic solvents, aprotic solvents, organic solvents, and any combination thereof. 25 13. The analytical method according to any one of claims 1–12, wherein said contacting is carried out in a solvent selected from chloroform, dichloromethane, acetonitrile, toluene, tetrahydrofuran, methanol, ethanol, isopropanol, dimethyl sulfoxide (DMSO), dimethylformamide (DMF), pentane, pentane isomers, hexane, hexane isomers, ether, 30 dichloroethane, acetone, ethyl acetate, butanone, and mixtures of any combination thereof. 310804994v3– 46 – 14. The analytical method according to any one of claims 1–13, wherein said contacting is carried out in the presence of triphenylphosphine (PPh3) and diisopropyl azodicarboxylate (DIAD). 5 15. The analytical method according to any one of claims 1–14, wherein said contacting is carried out under a nitrogen atmosphere.

16. The analytical method according to any one of claims 1–15, wherein said contacting is carried out for about 30 to about 200 minutes. 10 17. The analytical method according to any one of claims 1–16, wherein said contacting is carried out at ambient temperature.

18. The analytical method according to any one of claims 1–17, wherein the absolute 15 configuration of the chiral alcohol in the sample is determined.

19. The analytical method according to any one of claims 1–18, wherein the enantiomeric and / or diastereomeric composition of the chiral alcohol in the sample is determined. 20 20. The analytical method according to any one of claims 1–19, wherein both the absolute configuration of the chiral alcohol in the sample, and the enantiomeric and / or the diastereomeric composition of the chiral alcohol in the sample are determined.

21. The analytical method according to any one of claims 1-20, wherein said determining the 25 concentration of the chiral alcohol in the sample is carried out after determining the absolute configuration of the chiral alcohol in the sample, or the enantiomeric and / or the diastereomeric composition of the chiral alcohol in the sample, or both.

22. The analytical method according to claim 21, wherein said contacting is carried out in the 30 presence of a known amount of excess triphenylphosphine, wherein said determining the concentration of the chiral alcohol in the sample comprises: introducing a quinone compound into the reaction medium obtained following said contacting step to cause a reaction between triphenylphosphine and the quinone compound that produces a colorimetric change; and 35 determining the colorimetric change via UV-vis spectroscopy, wherein the colorimetric change is correlated to the amount of chiral alcohol in the sample. 310804994v3– 47 – 23. The analytical method according to any one of claims 21 or 22, wherein said contacting the sample is carried out on at least three measurements to which different known concentrations of the analyte are introduced; and said determining comprises generating a calibration curve, said generating comprising: 5 plotting recorded intensity measurements from the chiroptical assay (y-axis) versus the analyte concentration (x-axis) for the at least three measurements, and analyzing the plotted data using linear regression analysis.

24. The analytical method according to any one of claims 19 or 20, wherein said contacting the sample is carried out on at least three measurements to which different 10 known enantiomeric excesses of the analyte are introduced; and said determining comprises generating a calibration curve, said generating comprising: plotting recorded intensity measurements from the chiroptical assay (y-axis) versus the analyte enantiomeric excess (x-axis) for the at least three measurements, and analyzing the plotted data using linear regression analysis. 15 25. An aryl or heteroaryl sulfonamide probe having the structure according to formula (I): O O S1R2ArNRn (I), wherein Ar is a substituted or unsubstituted aromatic or heteroaromatic group, R1is H, a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl, 20 R2is a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl, and n is 0, 1, 2, 3, 4, or 5.

26. The aryl or heteroaryl sulfonamide probe according to claim 25, wherein the substituted or unsubstituted aryl at Ar, R1, and R2is independently selected from the group consisting of 25 phenyl, naphthyl, azulenyl, phenanthrenyl, anthracenyl, fluorenyl, pyrenyl, triphenylenyl, chrysenyl, naphthacenyl, biphenyl, triphenyl, and tetraphenyl.

27. The aryl or heteroaryl sulfonamide probe according to claim 25, wherein the substituted or unsubstituted heteroaryl at Ar, R1, and R2is independently selected from the group consisting 30 of pyrrolyl, pyrazolyl, imidazolyl, triazolyl, furyl, thiophenyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, triazinyl, thienopyrrolyl, furopyrrolyl, indolyl, azaindolyl, isoindolyl, indolinyl, indolizinyl, indazolyl, benzimidazolyl, imidazopyridinyl, benzotriazolyl, benzoxazolyl, benzoxadiazolyl, 310804994v3– 48 – benzothiazolyl, pyrazolopyridinyl, triazolopyridinyl, thienopyridinyl, benzothiadiazolyl, benzofuyl, benzothiophenyl, quinolinyl, isoquinolinyl, tetrahydroquinolyl, tetrahydroisoquinolyl, cinnolinyl, quinazolinyl, quinolizilinyl, phthalazinyl, benzotriazinyl, chromenyl, naphthyridinyl, acrydinyl, phenanzinyl, phenothiazinyl, phenoxazinyl, pteridinyl, and purinyl. 5 28. The aryl or heteroaryl sulfonamide probe according to claim 25, wherein Ar is selected from:wherein: 10 the squiggly line represents the point of attachment to the −S(O2)− group in formula I; each X is independently C or N, except that no more than three ring nitrogens are present in substituted or unsubstituted aromatic or heteroaromatic ring, and R3, R4, R5, R6, and R7are independently selected from the group consisting of a lone pair (when X is N), –H, –CN, –NO2, halogen, –C1-C6alkyl, –C1-C6alkoxy, –N–(alkyl)2, –C1-C615 alkenyl, –C1-C6alkynyl, – C1-C6perfluoroalkyl, –aryl, –perfluoroaryl, –aryloxy, –N–(aryl)2, –heteroaryl, –O–heteroaryl, –N–(heteroaryl)2, –cycloalkyl, –O–cycloalkyl, –N–(cycloalkyl)2, –heterocycloalkyl, –O–heterocycloalkyl, –N–(heterocycloalkyl)2, –C(O)Ra, –SO2Ra, and –OC(O)Ra; each Rais independently selected from the group consisting of –alkyl, –O–alkyl, –N–(alkyl)2, 20 –alkenyl, –alkynyl, –aryl, –O–aryl, –N–(aryl)2, –heteroaryl, –O–heteroaryl, –N–(heteroaryl)2, –cycloalkyl, –O–cycloalkyl, –N–(cycloalkyl)2, –heterocycloalkyl, –O–heterocycloalkyl, and –N–(heterocycloalkyl)2; and wherein, optionally, R3and R4, R4and R5, R5and R6, and / or R6and R7are alternatively taken together with the carbon or nitrogen atoms to which they are attached to form a fused 25 monocyclic or bicyclic ring system selected from the group consisting of cycloalkyl, heterocycloalkyl, aryl, and heteroaryl, wherein the fused ring system is optionally substituted with one or more groups selected from –alkyl, –O–alkyl, –N–(alkyl)2, – alkenyl, –alkynyl, –O-aryl, –O-heteroaryl, –N-(aryl)2, –N-(heteroaryl)2, –aryl, –C(O)Rc, –CO2Rb, –O-C(O)Rb, –NHC(O)Rb, –NRcC(O)Rb, –NO2, –CN, –halogen, and –SO2Rb, 30 wherein each Rbis independently aryl, heteroaryl, alkyl, or CH2Ar. 310804994v3– 49 – 29. The aryl or heteroaryl sulfonamide probe according to claim 25, wherein the probe is selected from:

530. A kit comprising: a solution comprising an aryl or heteroaryl sulfonamide probe according to any one of claims 25 to 29; and 10 optionally one or more of (i) sample tubes suitable for use with a spectrophotometer; (ii) an optically pure reference sample of a chiral alcohol; (iii) directions for using a spectrophotometer for carrying out circular dichroism (CD), vibrational CD (VCD), electronic CD, optical rotatory dispersion (ORD), or polarimetry 15 analyses to measure the concentration of an analyte in a sample and one or both of the absolute configuration of the analyte in the sample, and the enantiomeric and / or the diastereomeric composition of the analyte in the sample; and (iv) a recordable medium comprising a template for analyzing data obtained from the spectrophotometer and determining the concentration of an analyte in a sample and one or both 20 of the absolute configuration of the analyte in the sample, and the enantiomeric and / or the diastereomeric composition of the analyte in the sample.

31. The kit according to claim 30, wherein the probe is defined in any one of claims 26 to 28. 25 32. The kit according to claim 30, wherein the probe is selected from: 310804994v3– 50 –310804994v3

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