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.     

Determination of enantiomeric compositions of DOPA by tandem mass spectrometry using the kinetic method with fixed ligands

A fixed ligand (FL) version of the kinetic method was applied to rapid, simple, and accurate chiral analysis of DOPA, which is an important drug used for treatment of Parkinson's disease. Singly charged clusters containing the transition metal ion Cu(II), pyridyl ligands which serve as a fixed ligand, some amino acid as a reference, and the analyte DOPA were generated by electrospray ionization. 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(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 fixed ligands, references, and transition metals. Chiral discrimination was optimized in 1,10-phenanthroline as a FL, L-Phe and L-Pro as a reference, and Cu(II) as a central metal ion. Quantitative determinations of the enantiomeric composition of DOPA were achieved using two-point calibration curves. The linear relationship between the logarithm of the fragment ion abundance ratio (ln R) and enantiomeric compositions (ee%) of the DOPA allows the determination of the chiral purity of enantiomeric mixtures.     

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.     

Chiral and isomeric analysis by electrospray ionization and sonic spray ionization using the fixed-ligand kinetic method

The fixed-ligand version of the kinetic method has been used for chiral and for isomeric analysis by studying the dissociation kinetics of transition metal-bound trimeric cluster ions ([(M(II) + L(fixed)-H)(ref*)(An)](+), where M(II) is a transition metal, L fixed is a fixed (non-dissociating) ligand, ref* is a reference ligand and An is the analyte. The trimeric cluster ions are readily generated by electrospray ionization (ESI) or sonic spray ionization (SSI). The size of the fixed ligand, L- Phe-Gly-L-P he-Gly, is chosen based on previous results but with the inclusion of aromatic functionality to increase chiral recognition. Improved chiral/isomeric differentiation results from enhanced chiral/isomeric interactions (metal-ligand and ligand-ligand) due to the fixed ligand. As shown in the cases of chiral dipeptides (D-Ala-D-Ala/L-Ala-L-Ala), sugars (D/L-glucose, D/L-mannose) and isomeric tetrapeptides (L-Ala-Gly-Gly-Gly/Gly-Gly -Gly-L-Ala), improved chiral/isomeric discrimination by factors from three to six were obtained by the fixed ligand procedure. Chiral recognition is independent of the concentrations of the analyte, the reference ligand, the fixed ligand and the transition metal salt, a great advantage for practical applications. In addition to increased chiral distinction, the simplified dissociation kinetics also contribute to improved accuracy in chiral quantification, in comparison with data obtained by investigating the dissociation kinetics of simple trimeric cluster ions [M(II)(ref*)2(An) H](+). Accurate determination of enantiomeric excess (ee) is demonstrated by enantiomeric quantification of D-Ala-D-Ala/L-Ala-L-Ala down to 2% ee. Both ESI and SSI allow chiral quantification with similar accuracies. The performance of chiral analysis experiments is not limited to ion trapping devices such as quadrupole ion trap mass spectrometers by a hybrid quadrupole-time of flight (Q-ToF) mass spectrometer is shown to provide an alternative choice. The fixed-ligand kinetic method is not restricted to any particular kinds of isomers and, hence, represents a general procedure for improving molecular recognition and chiral analysis in the gas phase.     

Differentiation and quantitation of isomeric dipeptides by low-energy dissociation of copper(II)-bound complexes

Application of the kinetic method based on the dissociation of transition metal centered cluster ions is extended from chiral analysis (Tao, W. A.; Zhang, D.; Nikolaev, E. N.; Cooks, R. G. J. Am. Chem. Soc. 2000, 122, 10598) to quantitative analysis of isomeric mixtures, including those with Leu/Ile substitutions. Copper(II)-bound complexes of pairs of peptide isomers are generated by electrospray ionization mass spectrometry and the trimeric complex [CuII(ref)2(A) - H]+ (analyte A, a mixture of isomeric peptides; reference compound ref, usually a peptide) is caused to undergo collisional dissociation. Competitive loss of the neutral reference compound or the neutral analyte yields two ionic products and the ratio of rates of the two competitive dissociations, viz. the product ion branching ratio R is shown to depend strongly on the regiochemistry of the analyte in the precursor [CuII(A)(ref)2 - H]+ complex ion. Calibration curves are constructed by relating the branching ratio measured by the kinetic method, to the isomeric composition of the mixture to allow rapid quantitative isomer analysis.     

Halogen-substituted phenylalanines as enantioselective selectors for enantioselective discrimination of amino acids: effect of halogen

Halogen-substituted phenylalanines with a halogen X (X = F, Cl, Br or I) in the para position in the aromatic ring of L-phenylalanine are used as enantioselective selectors to explore the effect of the halogen substituent on the enantioselective discrimination of amino acids. Enantioselective discrimination is achieved by investigating the collision-induced dissociation spectra of the trimeric complex ion, [CuII(ref)2(A)-H]+, generated by electrospraying a solution of a mixture of D- or L-analyte amino acid (A), enantioselective reference ligand (ref) and CuCl2. The relative abundances of fragment ions resulting from the competitive loss of reference and analyte amino acids are considered for measuring the degree of enantioselective discrimination by applying the kinetic method. The enantioselectivity of the p-halogenated derivatives of L-Phe increases from fluorine to iodine for the studied amino acids (except for acidic amino acids). The validity of the present method has also been checked by cross enantioselective experiments using p-iodo-D-phenylalanine as the reference in place of p-iodo-L-phenylalanine. The enantioselectivity of fluoro-substituted L-phenylalanine is less than that obtained with L-phenylalanine. The high inductive effect of the fluorine atom decreases the strength of the pi-pi stacking interaction. The presence of halogen affects the enantioselectivity by inductive and steric effects.     

Chiral detection of entecavir stereoisomeric impurities through coordination with R‐besivance and ZnII using mass spectrometry

In this study, a mass spectrometry (MS)‐based kinetic method (KM) is shown to be successful at analyzing a multichiral center drug stereoisomer, entecavir (ETV), both qualitatively and quantitatively. On the basis of the KM, the bivalent complex ion [M^II(A)(ref*)₂]²⁺ (M^II = divalent metal ion, A = analyte, and ref* = chiral reference) was set as precursor ion in MS/MS. The experiment results suggest strong chiral selectivity between ETV and its isomers when using Zn^II coordinated with the chiral reference R‐besivance (R‐B). The logarithm of the fragment ion abundance ratio and the enantiomeric percentage (%) exhibits a strong linear relation because of the competitive loss of the reference and analyte. The product ion pair [Zn^II(R‐B)A‐H]⁺ (m/z 733) and [Zn^II(R‐B)₂‐H]⁺ (m/z 849), together with [R‐B + H]⁺ (m/z 394) and [A + H]⁺ (m/z 278), can realize the identification of ETV and all of its chiral isomers. Theoretical calculation were also performed using the B3LYP functional with the 6‐31G* and LanL2DZ basis set to clarify the mechanism of structural difference of these bivalent complex ions. The results reveal that MS‐KM can be used to detect optical impurities without a chiral chromatographic column and fussy sample pretreatment. The established method has been used to determine stereoisomeric impurities of less than 0.1% in ETV crude drug, a demonstration of its simple and effective nature for rapid detection of stereoisomeric impurities.     

PtCl

The ionic complex [PtCl(3)(C(2)H(4))](-)[(S,S)-(PhMeCH)(2)NH(2)](+) containing a chiral secondary amine is a new and versatile chiral derivatizing agent (CDA) for the determination of the enantiomeric composition of several unsaturated compounds including simple olefins: the diastereoisomeric mixtures arising from the exchange of ethylene by the unsaturated analytes are easily detected by (195)Pt NMR spectroscopy.     

Chiral Amplification of Oligopeptides in the Polymerization of α‐Amino Acid N‐Carboxyanhydrides in Water

This article is concerned with the chiral amplification of oligopeptides formed in the polymerization of chiral, nonracemic mixtures of the N‐carboxyanhydride (NCA) of Leu and Glu in aqueous solution. Labeling (deuteration) of one enantiomer and reversed‐phase and normal‐phase high‐performance liquid chromatography mass spectrometry (RP‐ and NP‐HPLC/MS, respectively) were used to determine the product distribution, both with respect to oligopeptide chain length and stereoisomer distribution. Starting the polymerization with an enantiomeric excess (ee) of 20% of the L ‐enantiomer (L ‐amino acid/D ‐amino acid 6 : 4) leads to an ee of 73% at the level of the homochiral enantiomeric (Leu)₅, and of 71% at the level of the homochiral enantiomeric (Glu)₇. For the Leu system and in the presence of a solid support (quartz), the ee reached values of up to 100%. We argue that such amplification processes could be relevant for the chemical evolution towards single‐handedness.     

Chiral discrimination of β‐3‐homo‐amino acids using the kinetic method

Chiral discrimination of seven enantiomeric pairs of β‐3‐homo‐amino acids was studied by using the kinetic method and trimeric metal‐bound complexes, with natural and unnatural α‐amino acids as chiral reference compounds and divalent metal ions (Cu²⁺ and Ni²⁺) as the center ions. The β‐3‐homo‐amino acids were selected for this study because, first of all, chiral discrimination of β‐amino acids has not been extensively studied by mass spectrometry. Moreover, these β‐3‐homo‐amino acids studied have different aromatic side chains. Thus, the emphasis was to study the effect of the side chain (electron density of the phenyl ring, as well as the difference between phenyl and benzyl side chains) for the chiral discrimination. The results showed that by the proper choice of a metal ion and a chiral reference compound, all seven enantiomeric pairs of β‐3‐homo‐amino acids could be differentiated. Moreover, it was noted that the β‐3‐homo‐amino acids with benzyl side chains provided higher enantioselectivity than the corresponding phenyl ones. However, increasing or decreasing the electron density of the aromatic ring by different substituents in both the phenyl and benzyl side chains had practically no role for chiral discrimination of β‐3‐homo‐amino acids studied. When copper was used as the central metal, the phenyl side chain containing reference molecules (S)‐2‐amino‐2‐phenylacetic acid (L ‐Phg) and (S)‐2‐amino‐2‐(4‐hydroxyphenyl)‐acetic acid (L ‐4′‐OHPhg) gave rise to an additional copper‐reduced dimeric fragment ion, [Cu^I(ref)(A)]⁺. The inclusion of this ion improved noticeably the enantioselectivity values obtained. Copyright © 2010 John Wiley & Sons, Ltd.     

Enantiomeric differentiation of β‐amino alcohols under electrospray ionization mass spectrometric conditions

The enantiomeric differentiation of a series of chiral β‐amino alcohols (A) is attempted, for the first time, by applying the kinetic method using L‐proline, L‐tryptophan, 4‐iodo‐L‐phenylalanine or 3, 5‐diiodo‐L‐tyrosine as the chiral references (Ref) and Cu²⁺ or Ni²⁺ ion (M) as the central metal ion. The trimeric diastereomeric adduct ions, [M+(Ref)₂+A‐H]⁺, formed under electrospray ionization conditions, are subjected for collision‐induced dissociation (CID) experiments. The products ions, formed by the loss of either a reference or an analyte, detected in the CID spectra are evaluated for the enantiomeric differentiation. All the references showed enantiomeric differentiation and the R_chiral values are better for the aromatic alcohols than for aliphatic alcohols. Notably, the R_chiral values of the aliphatic amino alcohols enhanced when Ni²⁺ is used as the central metal ion. The experimental results are well supported by computational studies carried out on the diastereomeric dimeric complexes. The computational data of amino alcohols is correlated with that of amino acids to understand the structural interaction of amino alcohols with reference molecule and central metal ion and their role on the stabilization of the dimeric complexes. Application of flow injection MS/MS method is also demonstrated for the enantiomeric differentiation of the amino alcohols. Copyright © 2014 John Wiley & Sons, Ltd.     

质谱动力学方法在手性分析中的应用

质谱动力学方法采用电喷雾离子化的方法获得含有手性化合物、并由过渡金属离子连接的复合物,然后进行碰撞诱导解离,两种解离途径的解离速率与混合物的对映体组成相关,进而可进行手性化合物光学纯度的测定。与其它常见手性测定方法相比,质谱动力学方法无需湿法化学处理和色谱分离步骤,操作简单、快速、灵敏,因而被广泛应用。本文对质谱动力学方法用于手性化合物对映体过量值测定的原理、改进方法、应用领域进行了评述。     

Colour indicator for enantiomeric excess and assignment of the configuration of the major enantiomer of an amino acid ester

A colour indicator for the full range of enantiomeric excess (-100%-->100% ee) is presented which is based on visual colour inspection of a liquid crystal doped with the analyte, i.e. the methyl ester of amino acid phenylglycine, providing the enantiomeric excess and allowing the assignment of the major enantiomer.     

Quantitative chiral analysis of phthaloylglutamic acid and related analogs by a single ratio kinetic method using electrospray ionization and matrix-assisted laser desorption techniques

A rapid method for quantitative chiral analysis of phthaloylglutamic acid and its dimethyl ester by Cook's kinetic method is demonstrated using electrospray ionization (ESI) and matrix-assisted laser desorption techniques. Transition-metal-bound complex ions containing the chiral phthaloylglutamic acid and its dimethyl ester are generated by ESI mass spectrometry and subjected to collision-induced dissociation. The ratio of the two competitive dissociation rates is related to the enantiomeric composition of the drug mixture. A seven-point calibration curve, derived from the kinetic method, allowed rapid quantitation of the enantiomeric excess of drug mixtures. In this paper, matrix-assisted laser desorption/ionization (MALDI) coupled with the linear ion trap (LIT) technique is evaluated for its applicability as a complementary technique to ESI for chiral discrimination and quantitation.     

Designing copper(II) ternary complexes to generate radical cations of peptides in the gas phase: role of the auxiliary ligand

A series of ternary copper(II) complexes of the type [Cu(II)(L)(M)](2+), where M represents the hexapeptides GGGFLR, YGGFLR and WGGFLR and L a set of 12 nitrogen donor ligands have been evaluated for their ability to form cationic peptide radicals, M(+)*, in the gas phase. Although the fragmentation chemistry of these ions is complex, two main conclusions emerge: (i) Complexes containing a tri- or tetra-dentate ligand were found to be more effective at producing the peptide radical because in these instances competitive loss of the ligand from the complex is inhibited; (ii) The ligands ought not possess any acidic protons in order to prevent competitive loss of the protonated peptide, [M + H](+). There is significant interaction of the N-terminal aromatic residues in YGGFLR and WGGLFR with the copper(ii) ion in several of the complexes as revealed by the formation of [Cu(I)(L)(p-quinomethide)](+) and [Cu(I)(L)(3-methyleneindoline)](+) fragment ions. Following its dissociation from the ternary complex, CID of the YGGFLR(+)* radical cation shows a dependence on the ligand in the complex from which it was formed. This 'memory effect' most likely reflects differences in the coordinated peptide structure induced by the ligand in the precursor complex which are maintained following dissociation.     

Nickel-Catalyzed Kinetic Resolution of Racemic Unactivated Alkenes via Enantio-, Diastereo-, and Regioselective Hydroamination

Kinetic resolution is a powerful strategy for the isolation of enantioenriched compounds from racemic mixtures, and the development of selective catalytic processes is an active area of research. Here, we present a nickel-catalyzed kinetic resolution of racemic α-substituted unconjugated carbonyl alkenes via the enantio-, diastereo-, and regioselective hydroamination. This protocol affords both chiral α-substituted butenamides and syn-β^2,3-amino acid derivatives with high enantiomeric purity (up to 99 % ee) and selectivity factor up to >684. The key to the excellent kinetic resolution efficiency is the distinctive architecture of the chiral nickel complex, which enables successful resolution and enantioselective C−N bond construction. Mechanistic investigations reveal that the unique structure of the chiral ligand facilitates a rapid migratory insertion step with one enantiomer. This strategy provides a practical and versatile approach to prepare a wide range of chiral compounds.     

Chiral recognition of zinc(II) ion complexes composed of bicyclo[3.3.0] octane-2,6-diol and <Emphasis Type="Italic">s</Emphasis>-Naproxen probed by collisional-induced dissociation

Chiral recognition of racemic bicyclo[3.3.0] octane-2,6-diol(B) was achieved in the gas phase using s-Naproxen(A) as reference, using the kinetics of competitive unimolecule dissociation of tetrameric zinc(II)-bound complexes by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometer(ESI-FTMS). As undergoing a mild competitive collision-induced dissociation(CID) experiment with a constant pressure argon gas introduced by leak valve, the tetrameric cluster ion [A(2)B(2)Z(n)(II)-H](+) forms only two trimeric ions and R(chiral) is subsequently obtained in the kinetic method. Further studies obtained the difference of Gibbs free energy of [ABZ(n)(II)-H](+)(Delta Delta G(ABZn(II)-H](+))) by dissociating [A(2)BZ(n)(II)-H](+), resulting two fragment ions [ABZ(n)(II)-H](+) and [A(2)Z(n)(II)-H](+), which can be established to a linear relationship between Delta Delta G([ABZn(II)-H](+)) and R(chiral)' basing on the kinetic method. The value of R(chiral)' suggested that Delta Delta G([ABZn(II)-H](+)) could be regarded as zero. Meanwhile, dissociation of [AB(2)Z(n)(II)-H](+) generated only one daughter ion [ABZ(n)(II)-H](+) in a stable pressure. Thus, a linear relationship was established between the difference of Gibbs free energy of [AB(2)Z(n)(II)-H](+)(Delta Delta G([AB(2)Zn(II)-H](+))) and R(chiral)" if the Delta Delta G([ABZn(II)-H](+)) can be negligible. Because there is also a linear relationship of R(chiral) in the tetrameric ion [A(2)B(2)Z(n)(II)-H](+) and the Gibbs energy difference of trimeric cluster ion [A(2)BZ(n)(+)(II)-H](Delta Delta G([A(2)BZn(II)-H](+))) plus that of [AB(2)Z(n)(II)-H](+), Delta Delta G([A(2)BZ(n)(II)-H]+]) is easy to be calculated in the dissociation process of tetrameric ion. Stable of R(chiral), R(chiral)' and R(chiral)" under different pressures show T(eff) does not affect the chiral recognition of cluster ions in the condition selected. If an only-one-daughter-ion fragment process of [A(2)BZ(n)(II)-H](+) was existed, R(chiral)' relating to this dissociation would be calculated just like R(chiral)" of [AB(2)Z(n)(II)-H](+) does. Conclusion was obtained that [A(2)BZ(n)(II)-H](+) makes more contribution to chiral recognition of tetrameric ion measured by kinetic method than [AB(2)Z(n)(II)-H](+) does as R(chiral)' and R(chiral)" were applied as index to evaluate the Gibbs free energy difference of these two trimeric cluster ions. Further discussion shows that steric interactions and pi-pi stacking interactions are the major factors responsible for the observed efficient chiral recognition in this system.     

Chiral discrimination of alpha-amino acids by the DNA triplet GCA using amino acids as a co-selector

The DNA triplet GCA is successfully used as a chiral selector for the chiral discrimination of amino acids using amino acids themselves as a co-selector. Chiral discrimination was achieved by investigating the collision-induced dissociation spectra of the [X(A) + X(R) + 2Y - 2H](2-) ion generated by electrospraying a mixture of analyte amino acid (X(A)), reference amino acid (X(R)) and GCA (Y). The relative abundances of fragment ions resulting from the competitive loss of reference and X(A)'s are considered for measuring the degree of chiral discrimination. GCA successfully shows D-selectivity for all the amino acids, except Tyr and Lys. The success of the method lies in the selection of a suitable 10(R) that has closer GCA binding affinity to that of analyte. The degree of discrimination by GCA is improved in the presence of the reference, and the chirality of the reference does not change the selectivity of GCA. The suitability of the method for the measurement of optical purity is also demonstrated.     

Multicomponent analyses of chiral samples by use of regression analysis of UV–visible spectra of cyclodextrin guest–host complexes

We report the first combined use of analytical spectroscopy, guest–host chemistry, and multivariate regression analysis for determination of enantiometric composition of multicomponent samples of chiral analytes. Sample solutions containing multicomponent analytes of ephedrine, tryptophan, propranolol, and proline of varying enantiomeric composition with beta-cyclodextrin (BCD) or methyl-beta-cyclodextrin (Me-BCD) as chiral host molecules were investigated using ultraviolet (UV)–visible spectroscopy. The interactions of enantiomers of chiral analytes with chiral hosts resulted in the formation of transient diastereomeric inclusion complexes with varying spectral properties. Multivariate analysis using partial-least-square (PLS) regression was used to correlate subtle changes in the UV–visible spectra of the guest–host complexes with the enantiomeric composition of the calibration samples. These PLS regressions were carefully optimized and then used to predict the enantiomeric composition of multicomponent chiral analytes of validation samples. The results of these validation studies demonstrate the predictive ability of the regression models for determination of future enantiomeric composition of samples. The accuracy of the models to correctly predict the enantiomeric composition of samples, evaluated by use of the root mean square percent relative error (RMS%RE) was analyte and chiral host dependent. In general, better prediction of enantiomeric composition of samples and low RMS%RE values were obtained when Me-BCD was used as the chiral host. The analyses procedure reported here is simple, rapid, and inexpensive. In addition, this approach does not require prior separation of chiral analytes, thus reducing analysis time and eliminating the need for expensive chiral columns.     

Enantiomer assays of amino acid derivatives using tertiary amine appended trans-4-hydroxyproline derivatives as chiral selectors in the gas phase

A pair of pseudo-enantiomers, tertiary amine appended trans-4-hydroxyproline derivatives were designed, synthesized, and evaluated as chiral selectors for enantiomer analysis of DNB-amino acid and their amides, in single-stage electrospray ionization/mass spectrometric experiments. The chiral selectors were designed to remove the interaction of the hydroxyl group of trans-4-hydroxyproline as well as separate the ionization site from the sites required for effective chiral recognition. Addition of a chiral analyte to a solution containing two pseudo-enantiomeric chiral selectors, affords selector-analyte complexes in the electrospray ionization mass spectrum where the ratio of these complexes is dependent on the enantiomeric composition of the analyte. The relationship between the ratio of the selector-analyte complexes in the electrospray ionization mass spectrum and the enantiomeric composition of the analyte can be used to relate the extent of the measured enantioselectivity and for quantitative enantiomeric composition determinations. Effects of acid modifiers (ammonium chloride, acetic acid, formic acid and hydrochloric acid) and instrument conditions on the selector-analyte ion intensity and the enantioselectivity (α_MS) were investigated. The largest α_MS was observed using ammonium chloride at a concentration around 0.5-1 mM at desolvation temperature of 150 °C. Capillary voltage has little effects on α_MS values. The sense of chiral recognition by MS is consistent with what is observed chromatographically. Quantitative enantiomeric composition determinations for N-(3,5-dinitrobenzoyl) leucinyl butylamide were performed. A comparison to the enantioselectivities towards a scope of analytes observed by chiral HPLC using a 3,5-dimethylanilide-proline-derived chiral stationary phase, is presented.