Chiral Reference Research Articles

Kinetic method for enantiomeric determination of thyroid hormone ( d, l-thyroxine) using electrospray ionization tandem mass spectrometry (ESI-MS/MS)

A rapid, sensitive, simple and accurate mass spectrometric analysis for the recognition and quantitation of d- and l-thyroxine ( d- and l-T 4) was achieved by using kinetic method. The method uses the kinetics of competitive unimolecular fragmentations of trimeric transition metal ion-bound clusters formed under electrospray ionization (ESI). Singly charged cluster ions containing the divalent central metal ion Ca(II)/Mn(II), an amino acid/modified amino acid chiral reference, and the analyte d- and l-T 4 were generated by ESI. The cluster ion of interest was mass-selected, and subjected to collision-induced dissociation for undergoing dissociation by competitive loss of either a neutral reference or a neutral analyte. The chiral selectivity ( R chiral), the ratio of the two competitive dissociation rates (abundances of fragment ion) containing the analyte in one enantiomeric form expressed relative to that for the fragments of the other enantiomer, ranges from 0.46 to 3.03. Method by using fixed ligand such as peptide has also successfully improved chiral recognition and quantitative accuracy, which simplifies the dissociation kinetics, in which only the reference ligand or the analyte can be lost. The linear relationship between the logarithm of the fragment ion abundance ratio (ln R) and enantiomeric compositions (ee%) of the T 4 allows the chiral purity of enantiomeric mixtures to be determined. The average relative errors were less than 2% between the actual and experimental enantiomeric compositions.

The use of X‐ray crystallography to determine absolute configuration

Essential background on the determination of absolute configuration by way of single-crystal X-ray diffraction (XRD) is presented. The use and limitations of an internal chiral reference are described. The physical model underlying the Flack parameter is explained. Absolute structure and absolute configuration are defined and their similarities and differences are highlighted. The necessary conditions on the Flack parameter for satisfactory absolute-structure determination are detailed. The symmetry and purity conditions for absolute-configuration determination are discussed. The physical basis of resonant scattering is briefly presented and the insights obtained from a complete derivation of a Bijvoet intensity ratio by way of the mean-square Friedel difference are exposed. The requirements on least-squares refinement are emphasized. The topics of right-handed axes, XRD intensity measurement, software, crystal-structure evaluation, errors in crystal structures, and compatibility of data in their relation to absolute-configuration determination are described. Characterization of the compounds and crystals by the physicochemical measurement of optical rotation, CD spectra, and enantioselective chromatography are presented. Some simple and some complex examples of absolute-configuration determination using combined XRD and CD measurements, using XRD and enantioselective chromatography, and in multiply-twinned crystals clarify the technique. The review concludes with comments on absolute-configuration determination from light-atom structures.

Chiral Discrimination of D- and L-Amino Acids Using Iodinated Tyrosines as Chiral References: Effect of Iodine Substituent

L-Tyrosine and iodinated L-tyrosines, i.e., 3-iodo-L-tyrosine and 3,5-diiodo-L-tyrosine, are successfully used as chiral references for the chiral discrimination of aliphatic, acidic, and aromatic amino acids. Chiral discrimination is achieved by investigating the collision-induced dissociation spectra of the trimeric complex [Cu(II)(ref)(2)(A) - H](+) ion generated by electro spraying the mixture of D- or L-analyte amino acid (A), chiral reference ligand (ref) and M(II)Cl(2) (M = Ni and Cu). The relative abundances of fragment ions resulted by the competitive loss of reference and analyte amino acids are considered for measuring the degree of chiral discrimination by applying the kinetic method. The chiral discrimination ability increases as the number of iodine atom increases on the aromatic ring of the reference and the discrimination is better with Cu when compared with Ni. A large chiral discrimination is obtained for aliphatic and aromatic amino acids using iodinated L-tyrosine as the reference. Computational studies on the different stabilities of the diastereomeric complexes also support the observed differences measured by the kinetic method. The suitability of the method in the measurement of enantiomeric excess over the range of 2% to 100% ee with relative error 0.28% to 1.6% is also demonstrated.

Enantiomer Separation of Amino Acids by Complexation with Chiral Reference Compounds and High-Field Asymmetric Waveform Ion Mobility Spectrometry: Preliminary Results and Possible Limitations

We present a new method for separation of enantiomers with high-field asymmetric waveform ion mobility spectrometry (FAIMS), coupled to mass spectrometric detection. Upon addition of an appropriate chiral reference compound to the analyte solution and subsequent ionization of the solution by electrospray ionization, analyte enantiomers formed diastereomeric complexes, which were potentially separable by FAIMS. The methodology being developed is intended to be general, but here amino acid analytes are specifically considered. In the examples presented herein, six pairs of amino acid enantiomers were successfully separated as metal-bound trimeric complexes of the form [MII(L-Ref)2(D/L-A)-H]+, where MII is a divalent metal ion, L-Ref is an amino acid in its L form acting as chiral reference compound, and A is the amino acid analyte. For example, D- and L-tryptophan were separated in FAIMS as [NiII(L-Asn)2(D-Trp)-H]+ and [NiII(L-Asn)2(L-Trp)-H]+. As FAIMS separation typically takes place over a time scale of only a few hundred milliseconds, the presented separation method opens new possibilities for rapid analysis of one analyte enantiomer in the presence of the other enantiomer. Preliminary quantification results are presented, which suggest that fast and sensitive quantitative chiral analyses can be performed with FAIMS. Method limitations are discussed in terms of diverse phenomena, which are not yet understood.

Improvements in quantitative chiral determinations using the mass spectrometric kinetic method

A significant shortcoming of the kinetic method for determining chiral purity from the competitive dissociation of a metal complex chelated with a chiral analyte and chiral reference is that it does not give useful quantitative data for high purity chiral samples in comparison to the standard chiral chromatography techniques. To extend the range of applications of the kinetic method to high chiral purity samples, a “vernier” method is reported which gives more accurate quantitative data for samples covering a narrow range of chiral purity. This is demonstrated for the case of a model amino acid in which Ni II is used as the metal cation, asparagine as the chiral reference, and the analyte is tryptophan. By selecting experimental conditions to give a measured peak abundance ratio near unity for the two competitive fragments (where abundance ratios are more accurately measured than at higher or lower values), samples of high (90–100%) chiral purity were more accurately assayed in the experiment using pure tryptophan. By switching the chirality of the reference compound, samples in the low (0–10%) chiral purity range could also be more precisely measured. The quantitative improvement in accuracy of the chiral measurement is shown by the average accuracy of 0.51% deviation (for the 90–100% chiral purity range of tryptophan using l-Asn as chiral reference) in comparison to the average deviation of 104.1% in the non-vernier region (the region that falls outside the high accuracy region of interest).

Stereoselective Discrimination and Quantification of Arginine and N-Blocked Arginine Enantiomers by Formation and Dissociation of Calcium-Mediated Diastereomeric Trimer Complexes with a Chiral Reference Compound Using Electrospray Ionization-Ion Trap Tandem Mass Spectrometry

Chiral resolution of arginine (Arg) and Arg derivatives is demonstrated using electrospray ionization-tandem mass spectrometry (ESI-MS). Calcium ion (Ca(II))-mediated trimeric clusters are generated, which incorporate the analyte of interest and an enantiomerically pure reference molecule of similar metal ion affinity. Two methods, one based on the measurement of a competitive-dissociation-based branching ratio (R(chir)) by the kinetic method (KM) and one based on the measurement of a chiral recognition ratio (CR) by a similar method, are compared. Incorporating N-blocked Arg derivatives (Z-Arg and Boc-Arg) as chiral references provides chiral resolution greater than that previously reported for Arg enantiomers. In a reciprocal manner, pure Arg enantiomers can be used as references for discriminating enantiomers of these N-blocked Arg derivatives. Condensed-phase and gas-phase Ca(II) ion affinity relative to Arg is also addressed qualitatively for other acidic, basic, and neutral amino acids. In some cases, when only one offspring ion is observed (insufficient for KM analysis), the CR method can be applied as an alternative to obtain a measurable stereoselectivity value for the system. The results of these experiments demonstrate the applicability of, and the difference between, the KM and the CR method for improved quantitative analysis of enantiomeric excess for Arg.

The complete chirospectroscopic signature of the peptide 3(10)-helix in aqueous solution.

We synthesized by solution methods a water-soluble, terminally blocked heptapeptide based on five markedly helicogenic, C(alpha)-tetrasubstituted alpha-amino acids C(alpha)-methyl-L-norvalines and two strongly hydrophilic 2-amino-3-[1-(1,4,7-triazacyclononane)]-L-propanoic acid residues at positions 2 and 5. A Fourier transform infrared absorption and NMR analysis in deuterated chloroform and aqueous solutions of the heptapeptide and two side-chain protected synthetic precursors confirmed our working hypothesis that all oligomers are folded in the 3(10)-helical conformation. Based on these findings, we exploited this heptapeptide as a chiral reference compound for detailed electronic CD, vibrational CD, and Raman optical activity characterizations of the 3(10)-helix in aqueous solution.

Chiral morphing and enantiomeric quantification in mixtures by mass spectrometry.

A novel mass spectrometric method is introduced for rapid and accurate chiral quantification by examining a tetracoordinated transition metal complex into which a reference and a fixed ligand are incorporated simultaneously with the analyte. Chiral analysis is performed by measuring the dissociation kinetics of these trimeric cluster ions [(M(II) + L(fixed) - H)(ref)(An)]+ (M(II) = a transition metal ion, L(fixed) = chiral peptide fixed ligand, ref = chiral reference ligand, and An = chiral analyte) in an ion trap mass spectrometer. The ratio of the product ion branching ratios measured when a pair of pure chiral fixed ligands and chiral reference ligands (/ref(D) and /ref(L); or /ref(L) and /ref(D)) are employed in separate experiments is related, via the kinetic method formalism, to the enantiomeric composition of the chiral mixture. This fixed-ligand quotient ratio (QR(fixed)) is logarithmically proportional to enantiomeric purity allowing construction of a calibration curve for chiral analysis when the analyte is only available in one form of known optical purity. There are reciprocal relationships when switching the chirality of the fixed/reference ligands. Improved quantification accuracy (due to simplified dissociation kinetics) and ready construction of two or more single-point calibration curves allow data to be cross-checked and represent an advantage of this approach. These features and the matrix tolerance of the kinetic method are demonstrated using the QR(fixed) method for determinations of enantiomeric excess of the drug DOPA in the presence of the co-drug compound L-carbidopa. The chiral selectivity of DOPA was found to vary from 0.0581 to 0.337 using this method, depending on the choices of fixed-ligand and reference chirality. The average relative errors are less than 1.2%. The potential of chiral morphing (changing chiral centers in the ligands) to further refine the chiral interactions and hence to maximize chiral recognition is shown.

Evaluation of chiral recognition characteristics of metal and proton complexes of di- o-benzoyl-tartaric acid dibutyl ester and L-tryptophan in the gas phase

Chiral recognition of di- o-benzoyl-tartaric acid dibutyl ester (T) was achieved in the gas phase by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry. In this method two divalent transition metal cations, zinc(II) and copper(II), were used as binding metal ions, and L-tryptophan (A) was used as a chiral reference. Multimeric complexes were readily formed by electrospray ionization of a methanol:water (50:50) solution containing metal ion, L-tryptophan and T. These multimeric complexes included singly charged protonated dimeric [TAH] +, doubly charged copper(II) bound tetrameric [TACu-H] 2 2+ and doubly charged zinc(II) bound tetrameric [TAZn-H] 2 2+, together with other complexes. The mass-selected complex, i.e., [TAH] +, [TACu-H] 2 2+ and [TAZn-H] 2 2+, was used to acquire the second stage mass spectra. The chiral recognition capability of these three complexes was evaluated using the abundance ratios of daughter ion to parent ion. A high degree of chiral recognition ability was observed in [TACu-H] 2 2+ and [TAZn-H] 2 2+. It was found that the type of binding ion played an important role in the chiral recognition. Different binding ions exhibited distinctive dissociation pathways and unique chiral recognition characteristics. The present method is based not only on whole-molecule loss but also on fractional-molecule loss. In addition, the reproducibility of the chiral recognition method was confirmed by several determinations of the abundance ratios of daughter ion to parent ion with a fixed activation energy and with five different activation energies. It was also shown that this chiral recognition method can tolerate acid interference.

Kinetic method for the simultaneous chiral analysis of different amino acids in mixtures.

The kinetic method has been extended to enantiomeric excess (ee) determinations on amino acids present in mixtures. Singly charged trimeric clusters [Cu(II)(ref*)(2)(A(m)) - H](+) are readily generated by electrospraying solutions containing Cu(II), a chiral reference ligand (ref*), and the amino acids (analytes A(m), m = 1-3). A trimeric cluster ion for each amino acid is individually mass-selected and then collisionally activated to cause dissociation by competitive loss of either the reference ligand or the analyte. For each analyte in the mixture, as shown from separate experiments, the logarithm of the ratio of the fragment abundances for the complex containing one enantiomer of this analyte expressed relative to that for the fragments of the corresponding complex containing the other enantiomer is linearly related to the enantiomeric composition of the amino acid. Formation and dissociation of each trimeric complex ion are shown to occur independently of the presence of other analytes. Chiral selectivity appears to be an intrinsic property and the chiral selectivity R(chiral(m)) measured from the mixture of analytes is equal to R(chiral) measured for the pure analyte. The sensitive nature of the methodology and the linear relationship between the logarithm of the fragment ion abundance ratio and the optical purity, characteristic of the kinetic method, allow the determination of chiral impurities of less than 2% ee in individual compounds present in mixtures by simply recording the ratios of fragment ion abundances in a tandem mass spectrum.

Chiral analysis using the kinetic method with optimized fixed ligands: applications to some antibiotics.

A new version of the kinetic method for chiral analysis, which employs a fixed (nondissociating) ligand as well as the usual analyte and chiral reference ligands, is introduced to simplify the kinetics of this experiment. Singly charged clusters containing the divalent transition metal ion MnII, a peptide which serves as a fixed ligand, an amino acid chiral reference, and the analyte 4-benzyl-2-oxazolidinone were generated by electrospray ionization (ESI). The cluster ion of interest was mass-selected, and the kinetics of its competitive unimolecular dissociations was investigated in an ion trap mass spectrometer. The chiral selectivity (R(fixed)chiral), the ratio of the two fragment ion abundances when the cluster contains one pure enantiomer of the analyte expressed relative to that for the other enantiomer, varies with increasing size of the fixed peptide ligands. The metal-ligand and the ligand-ligand interactions that produce chiral discrimination are optimized in the tetrapeptide fixed ligand Gly-Gly-Ala-Gly, as shown by data for 15 fixed ligands. The difference in the free energies of activation for the two competitive reactions is estimated to be approximately 7 kJ/mol for this particular fixed ligand. The sensitive nature of the methodology and the linear relationship between the logarithm of the fragment ion abundance ratio and the optical purity (intrinsic to the kinetic method) allows mixtures to be analyzed for as little as 1% enantiomeric excess (ee), by simply recording the ratios of fragment ion abundances in a tandem mass spectrum. These features are demonstrated in the case of the pharmacologically important 4-benzyl-2-oxazolidinones and in the case of penicillamine.

Ligand and metal-ion effects in metal-ion clusters used for chiral analysis of alpha-hydroxy acids by the kinetic method.

Chiral recognition of alpha-hydroxy acids has been achieved, and mixtures of enantiomers have been quantified in the gas phase, by using the kinetics of competitive unimolecular dissociation of singly-charged transition metal ion-bound trimeric complexes, [M(II)(A)(ref*)(2)-H](+) (M(II)=divalent transition metal ion; A=alpha-hydroxy acid; ref*=chiral reference ligand), to form the dimeric complexes [M(II)(A)(ref*)-H](+) and [M(II)(ref*)(2)-H](+). Chiral selectivity, the ratio of these two fragment ion abundances for the complex containing the analyte in one enantiomeric form expressed relative to that for the fragments of the corresponding complex containing the other enantiomer, ranges from 0.65 to 7.32. Chiral differentiation is highly dependent on the choice of chiral reference compound and central metal ion. The different coordination geometry of complexes resulting from the different d-orbital electronic configurations of these transition metal ions plays a role in chiral discrimination. Of all the transition metal ions examined chiral recognition is lowest for Cu(II), because of large distortion of the coordination complexes, and hence weak metal-ligand interactions and small stereochemical effects. It seems that two independent pi-cation interactions occur when N-acetyl-substituted aromatic amino acids used as the reference ligands and this accounts for improved chiral discrimination. If both metal-ligand and ligand-ligand interactions are optimized, large chiral selectivity is achieved. The sensitive nature of the methodology and the linear relationship between the logarithm of the fragment ion abundance ratio and the optical purity, which are intrinsic to the kinetic method, enable mixtures to be analyzed for small enantiomeric excess ( ee) by simply recording the ratios of fragment ion abundances in a tandem mass spectrum.

Quotient ratio method for quantitative enantiomeric determination by mass spectrometry.

The quotient ratio (QR) method, a new quantitative method for enantiomeric determination by mass spectrometry, is presented. The method is illustrated by the case in which a chiral analyte (A) and a chiral reference compound (ref*) are simultaneously coordinated to a transition metal ion (e.g., Cu2+ or Zn2 ) to form a singly charged deprotonated cluster ion in the course of electrospray ionization. The trimeric complex ion (three chiral ligands-2 mol of the analyte and 1 mol of the reference compound) is collisionally activated and undergoes dissociation by competitive loss of either a neutral reference or a neutral analyte. The ratio of the product ion branching ratios measured when pure chiral reference compounds (ref*D and ref*L) are employed in separate experiments is related via the kinetic method to the enantiomeric composition of the chiral mixture. This quotient ratio is logarithmically related to enantiomeric purity. In contrast to the previous single ratio method (Tao, W. A.; Zhang, D.; Wang, F.; Thomas, P. D.; Cooks, R. G. Anal. Chem. 1999, 71, 4427), the QR method reported here requires only one sample of the analyte with known optical purity to allow construction of a calibration curve. Accurate determinations of enantiomeric excess for amino acids and the chiral drug DOPA are demonstrated.

Quantitative chiral analysis of sugars by electrospray ionization tandem mass spectrometry using modified amino acids as chiral reference compounds.

Rapid quantitative enantiomeric analysis of mannose, glucose, galactose, and ribose is achieved using electrospray ionization and cluster ion dissociation with data analysis by the kinetic method. Several modified amino acids (N-Ac-L-Phe, N-benzoyl-L-Phe, N-t-Boc-L-Phe, N-Ac-L-Pro, N-t-Boc-L-Pro, N-Fmoc-L-Pro, N-Ac-L-Tyr, O-Me-L-Tyr) and four transition divalent metal cations (Co2+, Cu2+, Ni2+, and Zn2+) were tested to select the best system for chiral recognition and quantitation of each sugar. Quantitative determinations of the enantiomeric compositions of sugar solutions were achieved using either multiple- or two-point calibration curves; differences between the actual and experimental values were <2% enantiomeric excess (ee).

Mass spectrometric quantitation of chiral drugs by the kinetic method.

A novel mass spectrometric method for rapid, accurate (2-4% ee) quantitation of chiral drugs is described. Copper(II)-bound complexes of seven model drugs (atenolol, DOPA, ephedrine, pseudoephedrine, isoproterenol, norepinephrine, propranolol) with chiral reference compounds (L-amino acids) are generated by electrospray ionization mass spectrometry. The trimeric complex ions (three chiral ligands--one of the analyte and two of the reference compound) are collisionally activated, and they undergo dissociation by competitive loss of either the neutral reference or the neutral drug molecule. The ratio of the two competitive dissociation rates, viz. the product ion branching ratio, is related via the kinetic method to the enantiomeric composition of the drug mixture. A two-point calibration curve, derived from the kinetic method, allows rapid quantitation of enantiomeric excess of drug mixtures. The chiral sensitivity of the method is such as to allow determination of mixtures with a few percent enantiomeric contamination.

Chiral resolution of d- and l-amino acids by tandem mass spectrometry of Ni(II)-bound trimeric complexes

Mass-selected trimeric cluster ions, [Ni II(A)(ref∗) 2−H] +, where A designates the analyte amino acid and ref∗ designates the chiral reference amino acid, undergo competitive collision-induced dissociation to yield two dimeric clusters with a branching ratio dictated by the stereochemistry of both the analyte and the reference. This branching ratio ( R) is related, using the kinetic method, to an enthalpic term, ΔNi IIBDE, the difference between the formation enthalpies of the two dimeric clusters. Chiral resolution of d- and l-amino acids is directly related to the relative stabilities of the two diastereomeric clusters formed by the dissociation of the Ni(II)-bound trimeric clusters. The dimeric product ions differ in enthalpy by only a few kilojoules per mole, but chiral recognition is achieved for all 19 naturally occurring chiral amino acids, using appropriate reference amino acids. Because there is evidence that the dissociating trimeric clusters may exist in different isomeric forms, the proportion of analyte versus reference amino acids in the mixture was examined to study the effect of this ratio on the success of chiral recognition. The effect was found to be negligible. This suggests that there is an equilibrium between these isomeric clusters, which is governed by thermochemical properties rather than the relative concentrations of the constituent amino acids. A linear correlation was observed between ln( R) and enantiomeric compositions of the analyte as expected from the kinetic method treatment, and the direct measurement of optical purity to within 3% enantiomeric excess was demonstrated. With this new chiral recognition technique, qualitative and quantitative chiral analysis of amino acids is achieved.

Copper(II)-Assisted Enantiomeric Analysis of d,l-Amino Acids Using the Kinetic Method: Chiral Recognition and Quantification in the Gas Phase

Chiral recognition of d- and l-amino acids is achieved and mixtures of enantiomers quantified in the gas phase, using the kinetics of competitive unimolecular fragmentations of trimeric Cu(II)-bound complexes. Singly charged copper(II)−amino acid cluster ions [CuII(A)(ref*)2−H]+ (A = amino acid; ref* = chiral reference ligand, selected from among the natural α-amino acids) undergo competitive collision-induced dissociation (CID) in a quadrupole ion trap to form the dimeric complexes [CuII(A)(ref*)−H]+ and [CuII(ref*)2−H]+. The abundance ratio of these fragment ions depends strongly on the stereochemistry of the ligands in the precursor [CuII(A)(ref*)2−H]+ complex ion and specifically on the chirality of the analyte amino acid. The chiral selectivity, the ratio of the two fragment ion abundances for the complex containing one enantiomer of analyte expressed relative to that for the fragments of the corresponding complex containing the other enantiomer, ranges from 0.47 to 11. An energy quantity, Δ(ΔCuIIBDE), is predicted and shown to serve as a thermochemical indicator of chiral discrimination; its value is calculated from the fragment ion abundance ratios using the kinetic method of estimating thermochemical quantities from the kinetics of cluster ion dissociation. Large chiral distinctions are observed with all of the natural chiral α-amino acids, except cysteine and arginine, by appropriate choice of the reference ligand. The Δ(ΔCuIIBDE) values range from −2.2 to 6.9 kJ/mol. Amino acids with aromatic substituents display the largest chiral distinction, which is consistent with ligand exchange chromatographic results for analogous systems. The structures of the fragment Cu(II) complexes are discussed in the light of the CID behavior of related compounds. The interactions within these ions that might contribute to chiral recognition are rationalized to account for the observed chiral effects. The sensitive nature of the methodology and the linear relationship between the logarithm of the fragment ion abundance ratio and the optical purity, which is intrinsic to the kinetic method, allows mixtures to be analyzed for small enantiomeric excess (ee) by simply recording ratios of fragment ion abundances in a mass spectrum.

A highly functionalized ferrocenylpyrazolo[2,3-a]pyridine

.CommentIn view of the recent growing interest in the field of asym-metric synthesis, ferrocene and its derivatives occupy animportant place in synthetic and materials chemistry due totheir unique non-linear optical, liquid and crystalline andferromagnetic properties (Hayashi, 1995). The 1,2- and 1,3-disubstituted ferrocene derivatives, having planar chirality, aresuitable chiral reference systems for asymmetric inducedsynthesis (Marquarding et al., 1970). The unique properties ofthese compounds encouraged the synthesis of ferrocenederivatives with highly functionalized ring systems directlylinked to one of the cyclopentadiene moieties. Thesecompounds do not exhibit optical activity due to lack ofrestricted rotation. The title compound was synthesizedfollowing a procedure reported earlier (Nath et al., 1998). Thestructure of the title compound, (I), was assigned by IR, NMRand mass spectra, but the conformation of the ferrocenemoiety with respect to the heterocyclic ring was of particularinterest and prompted us to undertake the present X-raydiffraction study.The title molecule contains one pyrazolo[2,3-a]pyridinefused-ring system (PP), one bromo-phenyl ring (BP) and oneferrocenyl moiety. The two individual rings [inclined to eachother by an angle 1.9 (2)

Analysis of synthetic isoleucine conjugates of cucurbic acid isomers by liquid chromatography

Conjugates of 3,7-trans cucurbic acid isomers with either (S)- or (R)-isoleucine were synthesised from the diastereomeric conjugate of (3R, 7R)-jasmonic acid and (3S, 7S)-jasmonic acid by sodium borohydride reduction. The resulting diastereomers were characterised by nuclear magnetic resonance spectra. The authentic substances were subjected to liquid chromatography using a reversed-phase C18 matrix and the chiral stationary phase Chiralpak AS. For all (6RS)-hydroxy epimeric pairs, a baseline separation could be observed. The elution sequences obtained indicate that the 3,6-trans oriented epimers with (S)-isoleucine elute prior to the 3,6-cis configured individuals independent of the mode of chromatography. In contrast, the elution pattern of the conjugates containing (R)-isoleucine was reversed on the chiral stationary phase. The epimers with a 3,6-cis configured hydroxy group and the acid side chain eluted faster than those with the 3,6-trans configuration. The chromatographic conditions described are suitable for resolving mixtures of isomeric N-(cucurbinoyl)-isoleucines in order to assign their stereochemistry and to obtain chiral reference materials on a preparative scale. Copyright © 1999 John Wiley & Sons, Ltd.

Absolute Configuration of Chiral Ethanol-1-d: Neutron Diffraction Analysis of the (−)-(1S)-Camphanate Ester of (+)-(R)-Ethanol-1-d

The absolute configuration of (+)-ethanol-1-d has been determined to be R by the single-crystal neutron diffraction analysis of its (−)-camphanate ester. The absolute configuration of the (−)-camphanate group, which served as the chiral reference for the neutron study, was in turn established to be 1S,4R in an X-ray anomalous dispersion study of the complex Cu2(camphanate)4(ethanol)2. These results provide unambiguous confirmation that the optical rotation of (R)-ethanol-1-d (positive) is opposite to that of its higher homologs, (R)-propanol-1-d, (R)-butanol-1-d, and (R)-neopentanol-1-d (all negative), and demonstrates the usefulness of neutron diffraction in determining the absolute configuration of molecules possessing chiral methylene groups (i.e., molecules of the type CHDRR‘). Crystallographic details: for the neutron analysis of (+)-(R)-ethyl-1-d (−)-(1S)-camphanate: space group P21212 (orthorhombic), a = 6.422(1) A, b = 21.004(4) A, c = 9.275(2) A, V = 1251.1(7) A3, Z = 4; R(F2) = 0.083, and wR(F...